[ccp4bb] degradation of MBP fusion protein
Dear All: In my case, a 15KD protein without disufide bonds was expressed as inclusion bodies in E.coli but can be refolded as monomers with a very low solubility. Adding glycerol did not help so far. To increase the solubility, I fused my protein with maltose binding protein. After fusion, the protein can be solublly expressed but still in an aggregated form. There is also another problem. A large part of the MBP fusion protein was degraded somehow even at the condition of low temperature(18 degree) and low IPTG induction (0.1mM), which resulted in a truncated form of this protein, 30 out of 140 residues. However, this truncated form was monomeric. Based on this observation, I tried several short truncates as well, such as 80, 90, or 120 residues. But, after affinity purification, I got very similar results as mentioned above. I checked the sequence of my protein and there is no protease-sensitive site based on some protease-cutting-site-prediction servers. So, how can I deal with the degradation problem? and, how can I minimize the aggregation during expression? Any suggestions will be highly appreciated. Have a nice day! Jerry McCully By the way, thank folks for the answers of my another question about setting up view point in Pymol along axis in the unit cell.
Re: [ccp4bb] Reverse Translatase
I shell correct myself. The Darwin evolution of species is not sufficient to perform all functions of the reverse translatase. The Nature also uses viruses in order to "translate" proteins from different species. The other forms of reverse translation were probably not needed before the introduction of the immune system. Alex On Sep 7, 2010, at 7:26 PM, aaleshin wrote: Doesn't natural selection act like a Reverse Translatase? Which is quite an elegant implementation of the idea... On Sep 7, 2010, at 6:29 PM, Artem Evdokimov wrote: Regardless of whether a system like this exists in Nature or not - it's fun to imagine! On a microscopic scale one could propose a hypothetical mechanism by which a completely unfolded polypeptide chain could be fed into a gated (or state-locked) peptidase that may break the chain down in a co-ordinated stepwise fashion; releasing individual aa's into some sort of a nanoscale channel. The released aa's would then be sequentially coupled to something resembling tRNA - with pre-formed trinucleotides attached on the other end. Coupling would then presumably permit the triplets to ligate to one another sequentially - the resulting ssDNA or ssRNA would then have to be converted into a stable ds-form via the usual means, or otherwise protected in one of the usual ways. Codon space could be expanded by pre-loading carrier molecules with more than one type of triplet per carrier (biased towards whatever codon frequencies are prominent in the organism of choice) although this in no way resolves the random nature of the actual codon use within the resulting nucleotide sequence. The issue of amino acid coupling selectivity is pretty hairy - the best I could think of on a short notice is to have the receptor sites for individual aa's arranged in order of dropping selectivity -- however there is still the matter of shape/property similarities throwing wrenches into the works. An alternative would be a series of binary gates working on an exclusion principle. As to practicality of this kind of stuff - I am not sure; I can imagine an application similar to nano-scale multiparallel pyrosequencing: an unknown protein would be broken down into peptides via nonselective protease of some sort and then relatively short individual peptides are 'sequenced' in parallel, producing short DNA sequences that would later be complemented to dsDNA and allowed to cross-anneal and self-assemble via overlaps, similar to gapped gene assembly from short synthetic fragments (that first protease better be *really* non-specific!). At the end one could sequence the resulting long DNA to see what the original protein was like. A. On Tue, Sep 7, 2010 at 8:35 AM, David Schuller wrote: On 09/06/10 21:36, Jacob Keller wrote: Dear Crystallographers, does anyone know of any conceptual reason why a reverse translatase enzyme (protein-->nucleic acid) could not exist? I can think of so many things for which such an enzyme would be helpful, both to cells and to scientists...! Unless there is something I am missing, it would seem to me conceptually almost impossible that it *not* exist. See: "The RNA/Protein Symmetry Hypothesis: Experimental Support for Reverse Translation of Primitive Proteins" Masayuki Nahimoto, J. Theor. Biol. (2001) 209, pp 181-187. In which Nahimoto proposes such a system, and additionally proposes that it actually existed early in the development of life on this planet. Reasons why it "could not exist" - No. Reasons why it would be very difficult - yes. And plenty of reasons why Nahimoto is probably wrong about it having actually existed: There is absolutely no evidence presented that such a system was ever in operation in the history of life on this planet. Current theories such as the RNA World are much more likely explanations for how life as we currently know it may have developed from a pre- biotic state. DNA replication, DNA=>RNA transcription, and RNA=>Protein translation all depend on nucleic acid base pairing for part of their specificity. It truly is the secret of life. And it would not be especially helpful in Protein=>RNA reverse translation. Forward translation takes place in the ribosome, but extra specificity is "smuggled in" via a large set of tRNAs and tRNA charging enzymes, in reactions which took place beforehand, which are then made use of through the base-pairing codon:anti-codon recognition. Reverse translation would most definitely not be running forward translation in reverse; the specificity cannot be handled ahead of time, it needs to be available at the site of reverse translation itself when each successive peptide residue is presented. Progressivity: If different recognition sites are swapped in, this has to be done while keeping place in both the protein chain and in the growing nucleotide chain. Possibly the protein chain might be cleaved during the process. The chemistry and geometry of peptide residues is far more variable than tha
[ccp4bb] Increasing the resolution of protein
Dear all, I have a protein with molecular weight of 45 kDa which is part of a multisubunit complex. I was able to crystallize it, which diffracted to resolution of 8 Angstroms. I would like to increase the resolution of the protein crystals by trying to keep the protein intact with less possible truncations as its a part of a complex. I would be very happy to have suggestions in this concern. Thanks in advance. Regards, Ganesh
Re: [ccp4bb] Reverse Translatase
Doesn't natural selection act like a Reverse Translatase? Which is quite an elegant implementation of the idea... On Sep 7, 2010, at 6:29 PM, Artem Evdokimov wrote: Regardless of whether a system like this exists in Nature or not - it's fun to imagine! On a microscopic scale one could propose a hypothetical mechanism by which a completely unfolded polypeptide chain could be fed into a gated (or state-locked) peptidase that may break the chain down in a co-ordinated stepwise fashion; releasing individual aa's into some sort of a nanoscale channel. The released aa's would then be sequentially coupled to something resembling tRNA - with pre-formed trinucleotides attached on the other end. Coupling would then presumably permit the triplets to ligate to one another sequentially - the resulting ssDNA or ssRNA would then have to be converted into a stable ds-form via the usual means, or otherwise protected in one of the usual ways. Codon space could be expanded by pre-loading carrier molecules with more than one type of triplet per carrier (biased towards whatever codon frequencies are prominent in the organism of choice) although this in no way resolves the random nature of the actual codon use within the resulting nucleotide sequence. The issue of amino acid coupling selectivity is pretty hairy - the best I could think of on a short notice is to have the receptor sites for individual aa's arranged in order of dropping selectivity -- however there is still the matter of shape/property similarities throwing wrenches into the works. An alternative would be a series of binary gates working on an exclusion principle. As to practicality of this kind of stuff - I am not sure; I can imagine an application similar to nano-scale multiparallel pyrosequencing: an unknown protein would be broken down into peptides via nonselective protease of some sort and then relatively short individual peptides are 'sequenced' in parallel, producing short DNA sequences that would later be complemented to dsDNA and allowed to cross-anneal and self-assemble via overlaps, similar to gapped gene assembly from short synthetic fragments (that first protease better be *really* non-specific!). At the end one could sequence the resulting long DNA to see what the original protein was like. A. On Tue, Sep 7, 2010 at 8:35 AM, David Schuller wrote: On 09/06/10 21:36, Jacob Keller wrote: Dear Crystallographers, does anyone know of any conceptual reason why a reverse translatase enzyme (protein-->nucleic acid) could not exist? I can think of so many things for which such an enzyme would be helpful, both to cells and to scientists...! Unless there is something I am missing, it would seem to me conceptually almost impossible that it *not* exist. See: "The RNA/Protein Symmetry Hypothesis: Experimental Support for Reverse Translation of Primitive Proteins" Masayuki Nahimoto, J. Theor. Biol. (2001) 209, pp 181-187. In which Nahimoto proposes such a system, and additionally proposes that it actually existed early in the development of life on this planet. Reasons why it "could not exist" - No. Reasons why it would be very difficult - yes. And plenty of reasons why Nahimoto is probably wrong about it having actually existed: There is absolutely no evidence presented that such a system was ever in operation in the history of life on this planet. Current theories such as the RNA World are much more likely explanations for how life as we currently know it may have developed from a pre- biotic state. DNA replication, DNA=>RNA transcription, and RNA=>Protein translation all depend on nucleic acid base pairing for part of their specificity. It truly is the secret of life. And it would not be especially helpful in Protein=>RNA reverse translation. Forward translation takes place in the ribosome, but extra specificity is "smuggled in" via a large set of tRNAs and tRNA charging enzymes, in reactions which took place beforehand, which are then made use of through the base-pairing codon:anti-codon recognition. Reverse translation would most definitely not be running forward translation in reverse; the specificity cannot be handled ahead of time, it needs to be available at the site of reverse translation itself when each successive peptide residue is presented. Progressivity: If different recognition sites are swapped in, this has to be done while keeping place in both the protein chain and in the growing nucleotide chain. Possibly the protein chain might be cleaved during the process. The chemistry and geometry of peptide residues is far more variable than that of nucleotide residues. The genetic code of reverse translation would be completely independent of that in forward translation. For the two to have matched up (in the proposed naturally occurring RT system) would have been extremely fortuitous, imposing a strong barrier to the introduction of such a system. Difficulty in deal
Re: [ccp4bb] Reverse Translatase
In terms of "usefulness," I was actually thinking about cells learning how to make new proteins from other cells, or perhaps an immune system could use the info to make the right choice of starting materials. Also, codon bias could be explained as resulting from the nature of the reverse translatase machinery. Or an invader could copy the host's membrane proteins to evade detection. Ah, so many possibilities! And as I said before, considering that it would be so useful, and that the genius of macromolecular design observed in nature is apparently so unlimited, shouldn't it be out there somewhere? Nobel prize to the one who finds it... Jacob NB It should not cross our minds, I don't think, that if it were there, it would have been found. Small RNA phenomena, for example, went undetected for years, despite their commonness and high importance. - Original Message - From: "Artem Evdokimov" To: Sent: Tuesday, September 07, 2010 8:29 PM Subject: Re: [ccp4bb] Reverse Translatase Regardless of whether a system like this exists in Nature or not - it's fun to imagine! On a microscopic scale one could propose a hypothetical mechanism by which a completely unfolded polypeptide chain could be fed into a gated (or state-locked) peptidase that may break the chain down in a co-ordinated stepwise fashion; releasing individual aa's into some sort of a nanoscale channel. The released aa's would then be sequentially coupled to something resembling tRNA - with pre-formed trinucleotides attached on the other end. Coupling would then presumably permit the triplets to ligate to one another sequentially - the resulting ssDNA or ssRNA would then have to be converted into a stable ds-form via the usual means, or otherwise protected in one of the usual ways. Codon space could be expanded by pre-loading carrier molecules with more than one type of triplet per carrier (biased towards whatever codon frequencies are prominent in the organism of choice) although this in no way resolves the random nature of the actual codon use within the resulting nucleotide sequence. The issue of amino acid coupling selectivity is pretty hairy - the best I could think of on a short notice is to have the receptor sites for individual aa's arranged in order of dropping selectivity -- however there is still the matter of shape/property similarities throwing wrenches into the works. An alternative would be a series of binary gates working on an exclusion principle. As to practicality of this kind of stuff - I am not sure; I can imagine an application similar to nano-scale multiparallel pyrosequencing: an unknown protein would be broken down into peptides via nonselective protease of some sort and then relatively short individual peptides are 'sequenced' in parallel, producing short DNA sequences that would later be complemented to dsDNA and allowed to cross-anneal and self-assemble via overlaps, similar to gapped gene assembly from short synthetic fragments (that first protease better be *really* non-specific!). At the end one could sequence the resulting long DNA to see what the original protein was like. A. On Tue, Sep 7, 2010 at 8:35 AM, David Schuller wrote: On 09/06/10 21:36, Jacob Keller wrote: Dear Crystallographers, does anyone know of any conceptual reason why a reverse translatase enzyme (protein-->nucleic acid) could not exist? I can think of so many things for which such an enzyme would be helpful, both to cells and to scientists...! Unless there is something I am missing, it would seem to me conceptually almost impossible that it *not* exist. See: "The RNA/Protein Symmetry Hypothesis: Experimental Support for Reverse Translation of Primitive Proteins" Masayuki Nahimoto, J. Theor. Biol. (2001) 209, pp 181-187. In which Nahimoto proposes such a system, and additionally proposes that it actually existed early in the development of life on this planet. Reasons why it "could not exist" - No. Reasons why it would be very difficult - yes. And plenty of reasons why Nahimoto is probably wrong about it having actually existed: There is absolutely no evidence presented that such a system was ever in operation in the history of life on this planet. Current theories such as the RNA World are much more likely explanations for how life as we currently know it may have developed from a pre-biotic state. DNA replication, DNA=>RNA transcription, and RNA=>Protein translation all depend on nucleic acid base pairing for part of their specificity. It truly is the secret of life. And it would not be especially helpful in Protein=>RNA reverse translation. Forward translation takes place in the ribosome, but extra specificity is "smuggled in" via a large set of tRNAs and tRNA charging enzymes, in reactions which took place beforehand, which are then made use of through the base-pairing codon:anti-codon recognition. Reverse translation would most definitely not be running forward translation in reverse; the
Re: [ccp4bb] off topic-aggregation of proteins
Hi Bei, I have fought this problem myself several times. In addition to some of the suggestions from Tim, here are some suggestions for additives: As you mentioned Tween20 and Triton X-100 are very good for solubilizing difficult insoluble proteins. Those two are usually my first two weapons against insolubility. NP-40 (this is a detergent, but as far as I know it's not available commercially anymore, but you can buy "NP-40 substitute" from several companies) Glycerol (already mentioned) Xylitol or Sorbitol (which also work well as cryoprotectants if your protein/crystals like(s) them) Trehalose (+ or - extra phosphate in the buffer) 0.1M Arginine Also, it's best to start with low concentrations of these additives and increase them as needed. As for working with detergents, if your protein is soluble and not a membrane protein, you want to be well below the CMC. Like 50% of the CMC, max. The idea is that you want the soluble detergent molecules to interact with the sticky parts of your protein, and if the detergent molecules are forming micelles they aren't doing their job very effectively. Additionally, and I know this sounds a little insane - I was a skeptic at first, but others in my lab have had success with sonicating solutions of aggregated protein at very low intensity, very briefly. If your protein is reasonably stable but forms aggregates, I think this could work. (Like I said, I didn't beleive it at first, but CD scans of the sonicated protein show that it's still folded properly, and crystal structures have been solved from these samples.) One last thing to mention, when you add some of these additives to your buffers they will become contaminated very easily. Those stubborn little microorganisms that manage to contaminate buffers as simple as tris and salt will LOVE to eat up some of these sugars, amino acids, and even detergents, so be extra careful about autoclaving bottles, filtering buffers, etc. Good luck, as you are no doubt finding out this can be a difficult and frustrating problem. Mike Thompson - Original Message - From: "joybeiyang" To: CCP4BB@JISCMAIL.AC.UK Sent: Tuesday, September 7, 2010 12:30:04 PM GMT -08:00 US/Canada Pacific Subject: [ccp4bb] off topic-aggregation of proteins Hi everyone, sorry for this none CCP4 question, but I am working on some tough proteins which easily got aggregated after purification, I am thinking of using some detergents rightnow, however, I am a newbie in this field, could you please give me some advice on the usage of detergents? here is some basic information about the protein: 1. This protein is as big as 90KD. 2. The productivity of the protein is very low, and the homogeneity of the protein is not good. Most of the proteins come out of the column at void volumn, and the rest of the proteins forms 3-4 peaks, and each of them features low peak height. 3. I have already tried different truncates of the protein and homologous proteins from other species, up to now the protein on my hand is the best of them. 4. I have stocks of the following detergents:n-Octyl-b-D, n-Decyl-b-D, FOS-Choline-12, n-Dodecyl-b-D, Cymal-5, CHAPS, Tween20, Triton-X100. my question is: 1. would you please recommend some other detergents to try? 2. should I try different concentration of the detergent and how? right now I just tried the concentration equals the CMC, some of the detergents do improve the homogeneity of the protein, however, once I concentrate the protein for crystallization, the detergent get concentrated too, and as you know that is very bad for crystallization. 3. would you please recommend me some literature to resort to (about the same situation as mine or about the use of detergents)? 4. any other suggestions or comments about how to improve the quality of the protein. Your suggestions and comments will help me a lot and will be highly appreciated. Many thanks to all of you! Bei 2010-09-07 joybeiyang -- Michael C. Thompson Graduate Student Biochemistry & Molecular Biology Division Department of Chemistry & Biochemistry University of California, Los Angeles mi...@chem.ucla.edu
Re: [ccp4bb] Reverse Translatase
Regardless of whether a system like this exists in Nature or not - it's fun to imagine! On a microscopic scale one could propose a hypothetical mechanism by which a completely unfolded polypeptide chain could be fed into a gated (or state-locked) peptidase that may break the chain down in a co-ordinated stepwise fashion; releasing individual aa's into some sort of a nanoscale channel. The released aa's would then be sequentially coupled to something resembling tRNA - with pre-formed trinucleotides attached on the other end. Coupling would then presumably permit the triplets to ligate to one another sequentially - the resulting ssDNA or ssRNA would then have to be converted into a stable ds-form via the usual means, or otherwise protected in one of the usual ways. Codon space could be expanded by pre-loading carrier molecules with more than one type of triplet per carrier (biased towards whatever codon frequencies are prominent in the organism of choice) although this in no way resolves the random nature of the actual codon use within the resulting nucleotide sequence. The issue of amino acid coupling selectivity is pretty hairy - the best I could think of on a short notice is to have the receptor sites for individual aa's arranged in order of dropping selectivity -- however there is still the matter of shape/property similarities throwing wrenches into the works. An alternative would be a series of binary gates working on an exclusion principle. As to practicality of this kind of stuff - I am not sure; I can imagine an application similar to nano-scale multiparallel pyrosequencing: an unknown protein would be broken down into peptides via nonselective protease of some sort and then relatively short individual peptides are 'sequenced' in parallel, producing short DNA sequences that would later be complemented to dsDNA and allowed to cross-anneal and self-assemble via overlaps, similar to gapped gene assembly from short synthetic fragments (that first protease better be *really* non-specific!). At the end one could sequence the resulting long DNA to see what the original protein was like. A. On Tue, Sep 7, 2010 at 8:35 AM, David Schuller wrote: > On 09/06/10 21:36, Jacob Keller wrote: >> >> Dear Crystallographers, >> >> does anyone know of any conceptual reason why a reverse translatase enzyme >> (protein-->nucleic acid) could not exist? I can think of so many things >> for >> which such an enzyme would be helpful, both to cells and to scientists...! >> Unless there is something I am missing, it would seem to me conceptually >> almost impossible that it *not* exist. > > See: "The RNA/Protein Symmetry Hypothesis: Experimental Support for Reverse > Translation of Primitive Proteins" > Masayuki Nahimoto, J. Theor. Biol. (2001) 209, pp 181-187. > > In which Nahimoto proposes such a system, and additionally proposes that it > actually existed early in the development of life on this planet. > > Reasons why it "could not exist" - No. Reasons why it would be very > difficult - yes. And plenty of reasons why Nahimoto is probably wrong about > it having actually existed: > > There is absolutely no evidence presented that such a system was ever in > operation in the history of life on this planet. > > Current theories such as the RNA World are much more likely explanations for > how life as we currently know it may have developed from a pre-biotic state. > > DNA replication, DNA=>RNA transcription, and RNA=>Protein translation all > depend on nucleic acid base pairing for part of their specificity. It truly > is the secret of life. And it would not be especially helpful in > Protein=>RNA reverse translation. > > Forward translation takes place in the ribosome, but extra specificity is > "smuggled in" via a large set of tRNAs and tRNA charging enzymes, in > reactions which took place beforehand, which are then made use of through > the base-pairing codon:anti-codon recognition. > Reverse translation would most definitely not be running forward translation > in reverse; > the specificity cannot be handled ahead of time, it needs to be available at > the site of reverse translation itself when each successive peptide residue > is presented. > > Progressivity: If different recognition sites are swapped in, this has to be > done while keeping place in both the protein chain and in the growing > nucleotide chain. Possibly the protein chain might be cleaved during the > process. The chemistry and geometry of peptide residues is far more variable > than that of nucleotide residues. > > The genetic code of reverse translation would be completely independent of > that in forward translation. For the two to have matched up (in the proposed > naturally occurring RT system) would have been extremely fortuitous, > imposing a strong barrier to the introduction of such a system. > > Difficulty in dealing with post-translational modifications disulfides, > cyclical peptides, acetylation, phosphorylation, etc. > > A peptide sequ
Re: [ccp4bb] off topic-aggregation of proteins
Dear Bei, You question is not off topic, no apology needed. Have you considered other possibilities than using detergents? A little bit of glycerol might help, or e.g. a different salt, pH, buffer, some additive, or simply salt concentration. I once met a protein which was not soluble much beyond 1-2 mg/ml below 300mM NaCl and with 500mM NaCl could be concentrated to more than 100mg/ml. With the little amounts you have you could also try restricted proteolysis in order to determine stable subdomains. If you want to crystallise the protein, a detergent is not really favourable and it is worth playing with the above conditions before trying detergents. Tim On Tue, Sep 07, 2010 at 03:30:04PM -0400, joybeiyang wrote: > Hi everyone, sorry for this none CCP4 question, but I am working on some > tough proteins which easily got aggregated after purification, I am thinking > of using some detergents rightnow, however, I am a newbie in this field, > could you please give me some advice on the usage of detergents? > > here is some basic information about the protein: > 1. This protein is as big as 90KD. > 2. The productivity of the protein is very low, and the homogeneity of the > protein is not good. Most of the proteins come out of the column at void > volumn, and the rest of the proteins forms 3-4 peaks, and each of them > features low peak height. 3. I have already tried different truncates of the > protein and homologous proteins from other species, up to now the protein on > my hand is the best of them. 4. I have stocks of the following > detergents:n-Octyl-b-D, n-Decyl-b-D, FOS-Choline-12, n-Dodecyl-b-D, Cymal-5, > CHAPS, Tween20, Triton-X100. > > my question is: 1. would you please recommend some other detergents to try? > 2. should I try different concentration of the detergent and how? right now I > just tried the concentration equals the CMC, some of the detergents do improve > the homogeneity of the protein, however, once I concentrate the protein for > crystallization, the detergent get concentrated too, and as you know that is > very bad for crystallization. 3. would you please recommend me some > literature to resort to (about the same situation as mine or about the use of > detergents)? > 4. any other suggestions or comments about how to improve the quality of the > protein. > > Your suggestions and comments will help me a lot and will be highly > appreciated. > > Many thanks to all of you! > > Bei > > 2010-09-07 > > > > joybeiyang -- -- Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A signature.asc Description: Digital signature
[ccp4bb] off topic-aggregation of proteins
Hi everyone, sorry for this none CCP4 question, but I am working on some tough proteins which easily got aggregated after purification, I am thinking of using some detergents rightnow, however, I am a newbie in this field, could you please give me some advice on the usage of detergents? here is some basic information about the protein: 1. This protein is as big as 90KD. 2. The productivity of the protein is very low, and the homogeneity of the protein is not good. Most of the proteins come out of the column at void volumn, and the rest of the proteins forms 3-4 peaks, and each of them features low peak height. 3. I have already tried different truncates of the protein and homologous proteins from other species, up to now the protein on my hand is the best of them. 4. I have stocks of the following detergents:n-Octyl-b-D, n-Decyl-b-D, FOS-Choline-12, n-Dodecyl-b-D, Cymal-5, CHAPS, Tween20, Triton-X100. my question is: 1. would you please recommend some other detergents to try? 2. should I try different concentration of the detergent and how? right now I just tried the concentration equals the CMC, some of the detergents do improve the homogeneity of the protein, however, once I concentrate the protein for crystallization, the detergent get concentrated too, and as you know that is very bad for crystallization. 3. would you please recommend me some literature to resort to (about the same situation as mine or about the use of detergents)? 4. any other suggestions or comments about how to improve the quality of the protein. Your suggestions and comments will help me a lot and will be highly appreciated. Many thanks to all of you! Bei 2010-09-07 joybeiyang
Re: [ccp4bb] pyMol--set up view through one axis of the unit cell
Hi Jerry, If you wanted the mathematically exact axis, you _should_ be able to just make two pseudoatoms, position them at the proper position at the end of your axis and then use the orient command: pseudoatom axisMin, pos=[x1, y1, z1] pseudoatom axisMax, pos=[x2, y2, z2] orient axis* But, there is a bug with 'orient' on pseudoatoms: it's orienting the pseudoatoms along their 2nd principal axis, not the first. So, after you type those three command, follow up with: turn z, 90 and that should do it. Hope this helps, -- Jason On Tue, Sep 7, 2010 at 1:37 PM, Sampson, Jared wrote: > Hi Jerry, > > If your protein has an NCS symmetry axis parallel to a cell edge, you can try > using the “orient” command. > > http://www.pymolwiki.org/index.php/Orient > > Best, > Jared > > > On 9/3/10 7:31 PM, "James Stroud" wrote: > > On Sep 3, 2010, at 4:03 PM, Jerry McCully wrote: > It is a Pymol question. How can I set up the view through one axis of the > unit cell? > > By eye. Use orthoscopic view to help. Show the unit cell as a guide: > > http://www.pymolwiki.org/index.php/Cell > > James > > > > > -- > Jared Sampson > Xiangpeng Kong Lab > NYU Langone Medical Center > New York, NY 10016 > (212) 263-7898 > > > This email message, including any attachments, is for the sole use of the > intended recipient(s) and may contain information that is proprietary, > confidential, and exempt from disclosure under applicable law. Any > unauthorized review, use, disclosure, or distribution is prohibited. If you > have received this email in error please notify the sender by return email > and delete the original message. Please note, the recipient should check this > email and any attachments for the presence of viruses. The organization > accepts no liability for any damage caused by any virus transmitted by this > email. > = > -- Jason Vertrees, PhD PyMOL Product Manager Schrodinger, LLC (e) jason.vertr...@schrodinger.com (o) +1 (603) 374-7120
Re: [ccp4bb] pyMol--set up view through one axis of the unit cell
Hi Jerry, If your protein has an NCS symmetry axis parallel to a cell edge, you can try using the “orient” command. http://www.pymolwiki.org/index.php/Orient Best, Jared On 9/3/10 7:31 PM, "James Stroud" wrote: On Sep 3, 2010, at 4:03 PM, Jerry McCully wrote: It is a Pymol question. How can I set up the view through one axis of the unit cell? By eye. Use orthoscopic view to help. Show the unit cell as a guide: http://www.pymolwiki.org/index.php/Cell James -- Jared Sampson Xiangpeng Kong Lab NYU Langone Medical Center New York, NY 10016 (212) 263-7898 This email message, including any attachments, is for the sole use of the intended recipient(s) and may contain information that is proprietary, confidential, and exempt from disclosure under applicable law. Any unauthorized review, use, disclosure, or distribution is prohibited. If you have received this email in error please notify the sender by return email and delete the original message. Please note, the recipient should check this email and any attachments for the presence of viruses. The organization accepts no liability for any damage caused by any virus transmitted by this email. =
Re: [ccp4bb] Anybody using PVM by any chance ?
Fixed it, details are available if somebody is interested in them. Jürgen - Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry & Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Phone: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-3655 http://web.mac.com/bosch_lab/ On Sep 3, 2010, at 5:15 PM, Jürgen Bosch wrote: > Dear BB, > > I'm stuck with PVM (Parallel Virtual Machine) right now, trying to tell the > master to use a remote host as slave. > The individual machines run fine with PVM but they don't seem to communicate > with each other and I don't even get an error message, which makes it hard to > troubleshoot. > > If there is somebody on the BB with PVM experience, please contact me off the > board. > > Thanks in advance, > > Jürgen > > - > Jürgen Bosch > Johns Hopkins Bloomberg School of Public Health > Department of Biochemistry & Molecular Biology > Johns Hopkins Malaria Research Institute > 615 North Wolfe Street, W8708 > Baltimore, MD 21205 > Phone: +1-410-614-4742 > Lab: +1-410-614-4894 > Fax: +1-410-955-3655 > http://web.mac.com/bosch_lab/ >
[ccp4bb] Assistant Professor Vacancy
ASSISTANT PROFESSOR: The Department of Biochemistry and Cellular and Molecular Biology (BCMB) at the University of Tennessee, Knoxville seeks to fill a tenure-track faculty position at the assistant professor level to begin August 1, 2011 in the following area: EXPERIMENTAL PHYSICAL BIOCHEMIST Working in the area of nucleic acids or membranes. The successful applicant will be expected to develop a first-class, externally funded research program, to provide state-of-the-art training for graduate students and postdoctoral researchers, and to contribute to the teaching mission of the BCMB department at both the undergraduate and graduate levels. Required qualifications include a Ph.D. and postdoctoral experience in experimental molecular biophysics of nucleic acids, membranes, nucleic acid-protein or membrane-protein interactions, evidence of significant scientific productivity, and a commitment to an integrated program of teaching and research. Current research strengths of BCMB relevant to this search include Computational and Molecular Biophysics and Structural Biology. BCMB also has strength in Cell Biology, Genetics, Plant Biology, and Neurobiology. The successful candidate will benefit from interactions with strong research groups within UTK, access to supercomputing facilities (Kraken) and Computational and Physical Sciences programs at Oak Ridge National Laboratories (ORNL), which houses the new Spallation Neutron Source. The University welcomes and honors people of all races, creeds, cultures, and sexual orientations, and values intellectual curiosity, pursuit of knowledge, and academic freedom and integrity. Interested candidates should send a cover letter, a resume, a description of research experience and of the proposed research program, and arrange for three letters of reference to be sent to: Engin Serpersu, Chair, Faculty Search Committee, BCMB Department, M407 WLS, University of Tennessee, Knoxville, TN 37996-0840. Review of applications will begin on October 1, 2010 and will continue until the position is filled. All qualified applicants will receive equal consideration for employment and admissions without regard to race, color, national origin, religion, sex, pregnancy, marital status, sexual orientation, gender identity, age, physical or mental disability, or covered veteran status. Eligibility and other terms and conditions of employment benefits at The University of Tennessee are governed by laws and regulations of the State of Tennessee, and this non-discrimination statement is intended to be consistent with those laws and regulations. In accordance with the requirements of Title VI of the Civil Rights Act of 1964, Title IX of the Education Amendments of 1972, Section 504 of the Rehabilitation Act of 1973, and the Americans with Disabilities Act of 1990, The University of Tennessee affirmatively states that it does not discriminate on the basis of race, sex, or disability in its education programs and activities, and this policy extends to employment by the University. Inquiries and charges of violation of Title VI (race, color, national origin), Title IX (sex), Section 504 (disability), ADA (disability), Age Discrimination in Employment Act (age), sexual orientation, or veteran status should be directed to the Office of Equity and Diversity (OED), 1840 Melrose Avenue, Knoxville, TN 37996-3560, telephone (865) 974-2498 (V/TTY available) or 974-2440. Requests for accommodation of a disability should be directed to the ADA Coordinator at the Office of Equity and Diversity. Elias J. Fernandez, PhD Associate Professor University of Tennessee Biochemistry, Cellular & Molecular Biology 125 Austin Peay Knoxville, TN 37996 Phone:(865) 974-4090 Fax:(865) 974-6306 e-mail: elias.fernan...@utk.edu
Re: [ccp4bb] Reverse Translatase
On 09/06/10 21:36, Jacob Keller wrote: Dear Crystallographers, does anyone know of any conceptual reason why a reverse translatase enzyme (protein-->nucleic acid) could not exist? I can think of so many things for which such an enzyme would be helpful, both to cells and to scientists...! Unless there is something I am missing, it would seem to me conceptually almost impossible that it *not* exist. See: "The RNA/Protein Symmetry Hypothesis: Experimental Support for Reverse Translation of Primitive Proteins" Masayuki Nahimoto, J. Theor. Biol. (2001) 209, pp 181-187. In which Nahimoto proposes such a system, and additionally proposes that it actually existed early in the development of life on this planet. Reasons why it "could not exist" - No. Reasons why it would be very difficult - yes. And plenty of reasons why Nahimoto is probably wrong about it having actually existed: There is absolutely no evidence presented that such a system was ever in operation in the history of life on this planet. Current theories such as the RNA World are much more likely explanations for how life as we currently know it may have developed from a pre-biotic state. DNA replication, DNA=>RNA transcription, and RNA=>Protein translation all depend on nucleic acid base pairing for part of their specificity. It truly is the secret of life. And it would not be especially helpful in Protein=>RNA reverse translation. Forward translation takes place in the ribosome, but extra specificity is "smuggled in" via a large set of tRNAs and tRNA charging enzymes, in reactions which took place beforehand, which are then made use of through the base-pairing codon:anti-codon recognition. Reverse translation would most definitely not be running forward translation in reverse; the specificity cannot be handled ahead of time, it needs to be available at the site of reverse translation itself when each successive peptide residue is presented. Progressivity: If different recognition sites are swapped in, this has to be done while keeping place in both the protein chain and in the growing nucleotide chain. Possibly the protein chain might be cleaved during the process. The chemistry and geometry of peptide residues is far more variable than that of nucleotide residues. The genetic code of reverse translation would be completely independent of that in forward translation. For the two to have matched up (in the proposed naturally occurring RT system) would have been extremely fortuitous, imposing a strong barrier to the introduction of such a system. Difficulty in dealing with post-translational modifications disulfides, cyclical peptides, acetylation, phosphorylation, etc. A peptide sequencer coupled with a nucleotide synthesizer accomplishes somewhat the same thing, but on a macroscopic scale. This is an impediment to the motivation for constructing a reverse translatase enzymatic system. Cheers, -- === All Things Serve the Beam === David J. Schuller modern man in a post-modern world MacCHESS, Cornell University schul...@cornell.edu
Re: [ccp4bb] Fd-3m with pdbset
Dear Kevin (and others), The symmetry operators are in the "official" pdb in REMARK 290 records, both as text (e.g. x,y,z) and as symmetry (SMTRY) matrices and are even mandatory (see extract below). So far they just have been ignored by the CCP4 and most other crystallographic programs. If the CCP4 would agree to use these matrices or operators, the chaos with space group names and numbers would be solved or at least would not matter any more since they would not be used to get symmetry information. Best regards, Herman REMARK 290, Crystallographic Symmetry Remark 290 is mandatory for crystalline studies. The remark is generated by PDB. Example 1 2 3 4 5 6 7 1234567890123456789012345678901234567890123456789012345678901234567890 REMARK 290 REMARK 290 CRYSTALLOGRAPHIC SYMMETRY REMARK 290 SYMMETRY OPERATORS FOR SPACE GROUP: P 21 21 21 REMARK 290 REMARK 290 SYMOP SYMMETRY REMARK 290 NNNMMM OPERATOR REMARK 290 1555 X,Y,Z REMARK 290 2555 1/2-X,-Y,1/2+Z REMARK 290 3555 -X,1/2+Y,1/2-Z REMARK 290 4555 1/2+X,1/2-Y,-Z REMARK 290 REMARK 290 WHERE NNN -> OPERATOR NUMBER REMARK 290 MMM -> TRANSLATION VECTOR REMARK 290 REMARK 290 CRYSTALLOGRAPHIC SYMMETRY TRANSFORMATIONS REMARK 290 THE FOLLOWING TRANSFORMATIONS OPERATE ON THE ATOM/HETATM REMARK 290 RECORDS IN THIS ENTRY TO PRODUCE CRYSTALLOGRAPHICALLY REMARK 290 RELATED MOLECULES. REMARK 290 SMTRY1 1 1.00 0.00 0.00 0.0 REMARK 290 SMTRY2 1 0.00 1.00 0.00 0.0 REMARK 290 SMTRY3 1 0.00 0.00 1.00 0.0 REMARK 290 SMTRY1 2 -1.00 0.00 0.00 36.30027 REMARK 290 SMTRY2 2 0.00 -1.00 0.00 0.0 REMARK 290 SMTRY3 2 0.00 0.00 1.00 59.50256 REMARK 290 SMTRY1 3 -1.00 0.00 0.00 0.0 REMARK 290 SMTRY2 3 0.00 1.00 0.00 46.45545 REMARK 290 SMTRY3 3 0.00 0.00 -1.00 59.50256 REMARK 290 SMTRY1 4 1.00 0.00 0.00 36.30027 REMARK 290 SMTRY2 4 0.00 -1.00 0.00 46.45545 REMARK 290 SMTRY3 4 0.00 0.00 -1.00 0.0 REMARK 290 REMARK 290 REMARK: -Original Message- From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Kevin Cowtan Sent: Tuesday, September 07, 2010 12:54 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Fd-3m with pdbset George M. Sheldrick wrote: > Perhaps I should mention that with the SHELX method of specifying the > space group symmetry using the symmetry operators, alternative > settings, specified in IT or not, cause no problems. But then we would > not have had this thread (or the H3/R3 and P21221 threads). What George said. (This is the approach used by the CCP4 C libraries and clipper for mtzs and maps, and also for the coot-CNS interchange format, but it is not used for PDB files.) Is anyone who is involved in the new PDB format willing to campaign for the compulsory inclusion of the symops (either full set or a minimal set of generator ops) in the PDB file as a canonical representation of the spacegroup? -- EMAIL DISCLAIMER http://www.york.ac.uk/docs/disclaimer/email.htm
Re: [ccp4bb] Fd-3m with pdbset
George M. Sheldrick wrote: Perhaps I should mention that with the SHELX method of specifying the space group symmetry using the symmetry operators, alternative settings, specified in IT or not, cause no problems. But then we would not have had this thread (or the H3/R3 and P21221 threads). What George said. (This is the approach used by the CCP4 C libraries and clipper for mtzs and maps, and also for the coot-CNS interchange format, but it is not used for PDB files.) Is anyone who is involved in the new PDB format willing to campaign for the compulsory inclusion of the symops (either full set or a minimal set of generator ops) in the PDB file as a canonical representation of the spacegroup? -- EMAIL DISCLAIMER http://www.york.ac.uk/docs/disclaimer/email.htm
[ccp4bb] Job for scientific programmer, for Electron microscopy applications
Enquiries to Alan Roseman, please. m > Scientific Programmer, for Electron microscopy applications: > Faculty of Life Sciences, University of Manchester, UK > > Closing date: 14/09/2010, Reference: LS/10769 > > We seek a scientific programmer with an interest in structural biology > to join Dr Alan Roseman's group. Single particle analysis using > electron microscopy is a powerful and versatile technique for > determining molecular structures, and near atomic resolutions have > been be reached with some specimens. We are pioneering new methods for > analysis of structures of macromolecular complexes using electron > cryo-microscopy, making use of novel correlation functions and network > analysis. > > You will develop and program new software applications and algorithms, > and field test these on laboratory projects. Expertise in high > performance computing, and programming in the Fortran 90 and python > languages is required. > > You will have degree in computer science, computational biology, > engineering, or a related field (or equivalent experience). > > The post is funded by the BBSRC, starting from 1 November 2010 for up > to 15 months. > Salary > > £24,273 - £27,319 p.a. > Informal enquiries > > Dr Alan Roseman > Tel: + 44 (0) 161 275 7226 > Email alan.rose...@manchester.ac.uk -- *** * * * Dr. Martyn Winn * * * * STFC Daresbury Laboratory, Daresbury, Warrington, WA4 4AD, U.K. * * Tel: +44 1925 603455E-mail: martyn.w...@stfc.ac.uk* * Fax: +44 1925 603634Skype name: martyn.winn * * URL: http://www.ccp4.ac.uk/martyn/ * ***
[ccp4bb] ARCIMBOLDO - successful solution
Dear, Because this can certainly be of interest to other crystallographers, I would like to post the following positive comment: The recently developed software ARCIMBOLDO: Crystallographic Ab Initio protein solution far below atomic resolution, (Rodriguez et al., Nature Methods, 6, 651, 2009) http://chango.ibmb.csic.es/ARCIMBOLDO/ was able to find a successful solution for one of our protein structures. The protein showed only 30% sequence identity and ,although we've built several homology models, failed all MR attempts. Furthermore, crystal data were complete (1.6 angstrom), showed high redundancy (cubic space group) and lot's of alpha helices. In the future, a public web server will be provided to run the software on, but in the meantime very kind help was provided by the authors Dayté Rodriguez and Isabel Uson. I hope this comment will stimulate the use and the further development/implementation of the program. Regards Kristof -- Kristof Van Hecke, PhD Biomoleculaire Architectuur Celestijnenlaan 200 F B-3001 Heverlee (Leuven) Tel: +32(0)16327477 --
Re: [ccp4bb] Reverse Translatase
Whoops, sorry of course that is right. But 3 amino acids would be 8000, etc. -Greg On Sep 7, 2010, at 12:46 AM, Murray, James W wrote: So, let's say instead these adaptors recognize 2 amino acids at a time (still probably not robust enough). Then, one would need 2^20 adaptors, already a far greater number of gene products than that present in any genome than I know of... Surely only 20^2, which is 400? A lot, but managable. James -- Dr. James W. Murray David Phillips Research Fellow Division on Molecular Biosciences Imperial College, LONDON Tel: +44 (0)20 759 48895 From: CCP4 bulletin board [ccp...@jiscmail.ac.uk] On Behalf Of Greg Alushin [galus...@berkeley.edu] Sent: Tuesday, September 07, 2010 3:19 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Reverse Translatase Hi Jacob- What an intriguing proposition. I can think of multiple reasons why such a system would not exist, but there is a mechanistic one which is most fundamental, having to do with the nature of the genetic code. Say that there is a cellular machine which would unfold a protein and expose it to some sort of reading system (already a hard problem). There is now the issue of transforming the amino acid information into nucleic acid information. For simplicity let's assume that this system only uses one codon per amino acid, bypassing the degeneracy problem. How would the cell then read off the amino acid sequence? It seems the simplest solution would be analogous to translation, i.e. having adaptor molecules analogues to tRNAs which would guide an enzyme that synthesized the nucleic acid. Otherwise, one would have to invoke the idea of a single enzyme recognizing every amino acid, which seems impossible to me. As we know, the problem of protein-protein recognition is relatively complex. At a minimum, one would need 20 adaptor proteins to recognize the 20 canonical amino acids: however, it seems unlikely that recognition of a single amino acid would be robust enough to select for the correct adaptor molecule. So, let's say instead these adaptors recognize 2 amino acids at a time (still probably not robust enough). Then, one would need 2^20 adaptors, already a far greater number of gene products than that present in any genome than I know of... It might be tempting to draw an analogy between this system and the immune system, where an incredible diversity is generated from a small number of genes. However, diverse immune proteins all take the same input sequence (say antigen recognition) and lead to a single response, whereas this system has a 1 to 1 correspondence between inputs (protein sequence) and outputs (nucleic acid sequences), and thus there is no way that a randomization system could generate the required diversity. Cheers, -Greg Alushin Nogales lab UC Berkeley On Sep 6, 2010, at 7:12 PM, Michael Thompson wrote: Jacob, The idea is enticing, but don't forget that there are multiple degenerate codons for a given amino acid. Once the protein is synthesized, the specific codon information is lost. I think that's a fundamental problem. Keep the ideas coming, Mike Thompson - Original Message - From: "Jacob Keller" To: CCP4BB@JISCMAIL.AC.UK Sent: Monday, September 6, 2010 6:36:14 PM GMT -08:00 US/Canada Pacific Subject: [ccp4bb] Reverse Translatase Dear Crystallographers, does anyone know of any conceptual reason why a reverse translatase enzyme (protein-->nucleic acid) could not exist? I can think of so many things for which such an enzyme would be helpful, both to cells and to scientists...! Unless there is something I am missing, it would seem to me conceptually almost impossible that it *not* exist. Best Regards, Jacob Keller *** Jacob Pearson Keller Northwestern University Medical Scientist Training Program Dallos Laboratory F. Searle 1-240 2240 Campus Drive Evanston IL 60208 lab: 847.491.2438 cel: 773.608.9185 email: j-kell...@northwestern.edu *** -- Michael C. Thompson Graduate Student Biochemistry & Molecular Biology Division Department of Chemistry & Biochemistry University of California, Los Angeles mi...@chem.ucla.edu
Re: [ccp4bb] Reverse Translatase
>So, let's say instead these adaptors recognize 2 amino acids at a time >(still probably not robust enough). Then, one would need 2^20 >adaptors, already a far greater number of gene products than that >present in any genome than I know of... Surely only 20^2, which is 400? A lot, but managable. James -- Dr. James W. Murray David Phillips Research Fellow Division on Molecular Biosciences Imperial College, LONDON Tel: +44 (0)20 759 48895 From: CCP4 bulletin board [ccp...@jiscmail.ac.uk] On Behalf Of Greg Alushin [galus...@berkeley.edu] Sent: Tuesday, September 07, 2010 3:19 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Reverse Translatase Hi Jacob- What an intriguing proposition. I can think of multiple reasons why such a system would not exist, but there is a mechanistic one which is most fundamental, having to do with the nature of the genetic code. Say that there is a cellular machine which would unfold a protein and expose it to some sort of reading system (already a hard problem). There is now the issue of transforming the amino acid information into nucleic acid information. For simplicity let's assume that this system only uses one codon per amino acid, bypassing the degeneracy problem. How would the cell then read off the amino acid sequence? It seems the simplest solution would be analogous to translation, i.e. having adaptor molecules analogues to tRNAs which would guide an enzyme that synthesized the nucleic acid. Otherwise, one would have to invoke the idea of a single enzyme recognizing every amino acid, which seems impossible to me. As we know, the problem of protein-protein recognition is relatively complex. At a minimum, one would need 20 adaptor proteins to recognize the 20 canonical amino acids: however, it seems unlikely that recognition of a single amino acid would be robust enough to select for the correct adaptor molecule. So, let's say instead these adaptors recognize 2 amino acids at a time (still probably not robust enough). Then, one would need 2^20 adaptors, already a far greater number of gene products than that present in any genome than I know of... It might be tempting to draw an analogy between this system and the immune system, where an incredible diversity is generated from a small number of genes. However, diverse immune proteins all take the same input sequence (say antigen recognition) and lead to a single response, whereas this system has a 1 to 1 correspondence between inputs (protein sequence) and outputs (nucleic acid sequences), and thus there is no way that a randomization system could generate the required diversity. Cheers, -Greg Alushin Nogales lab UC Berkeley On Sep 6, 2010, at 7:12 PM, Michael Thompson wrote: > Jacob, > > The idea is enticing, but don't forget that there are multiple > degenerate codons for a given amino acid. Once the protein is > synthesized, the specific codon information is lost. > > I think that's a fundamental problem. > > Keep the ideas coming, > > Mike Thompson > > > > > - Original Message - > From: "Jacob Keller" > To: CCP4BB@JISCMAIL.AC.UK > Sent: Monday, September 6, 2010 6:36:14 PM GMT -08:00 US/Canada > Pacific > Subject: [ccp4bb] Reverse Translatase > > Dear Crystallographers, > > does anyone know of any conceptual reason why a reverse translatase > enzyme > (protein-->nucleic acid) could not exist? I can think of so many > things for > which such an enzyme would be helpful, both to cells and to > scientists...! > Unless there is something I am missing, it would seem to me > conceptually > almost impossible that it *not* exist. > > Best Regards, > > Jacob Keller > > > *** > Jacob Pearson Keller > Northwestern University > Medical Scientist Training Program > Dallos Laboratory > F. Searle 1-240 > 2240 Campus Drive > Evanston IL 60208 > lab: 847.491.2438 > cel: 773.608.9185 > email: j-kell...@northwestern.edu > *** > > -- > Michael C. Thompson > > Graduate Student > > Biochemistry & Molecular Biology Division > > Department of Chemistry & Biochemistry > > University of California, Los Angeles > > mi...@chem.ucla.edu
Re: [ccp4bb] Fd-3m with pdbset - test pdb file
Dear Herman, thanks for the PDB-file. I can confirm that there are no problems with this file at all. Paul has been in contact with me to work on the problem. Tim On Tue, Sep 07, 2010 at 08:57:49AM +0200, herman.schreu...@sanofi-aventis.com wrote: > Dear bulletin board, > > Tim asked me for the pdb file I used for testing (coot version 0.6), > which I have attached here. It is a totally artifical pdb file with one > random small ligand in it. Using a symmetry radius of 50 Angstrom, one > can get an idea of the symmetry in 'F d -3 m'. If in Tim's case with a > real protein coot comes to a grinding halt, it looks to me more like a > question of memory management than a problem with the symmetry > definitions. > > Best, > Herman > > -Original Message- > From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of > herman.schreu...@sanofi-aventis.com > Sent: Monday, September 06, 2010 2:44 PM > To: CCP4BB@JISCMAIL.AC.UK > Subject: Re: [ccp4bb] Fd-3m with pdbset > > Hi Tim, > > The string 'Fd-3m:1' is recognized by pdbset, but still produces 'F d -3 > m :'. I did a test with a single small ligand with coot and if you > delete the ':' from the CRYST1 card to make 'F d -3 m' coot happily > accepts it and seems to produce the correct symmetry mates. I hope that > this way you are able to display your molecule. I agree that there must > be a bug somewhere. > > Best, > Herman > > -Original Message- > From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of > Tim Gruene > Sent: Monday, September 06, 2010 1:38 PM > To: CCP4BB@JISCMAIL.AC.UK > Subject: [ccp4bb] Fd-3m with pdbset > > Hello, > I tried to assign a cell with space group F d -3 m to a PDB file using > pdbset. > Since it did not recognize the string 'F d -3 m' (nor 'F d -3 m :1') , I > used the space group number 227. This resulted in the space group string > 'F d -3 m :' > in the PDB-file instead of 'F d -3 m :1' as it is listed in syminfo.lib, > and hence coot refused to display the symmetry mates for the molecule. > > When I add the missing '1' in the resulting PDB-file, coot does display > the cell, but when I ask to display the symmetry mates, coot chokes and > consumed all my cpu power (for 8 molecules present in the unit cell...) > until I killed it. > > I was wondering if this is a known bug in syminfo.lib (and/or the > dependent > programs) and if there is a way around it (without using > phenix.get_me_out_of_here, I could of course use xp instead of coot for > such a small molecule). > > Kind regards, Tim > > P.S.: The bug in pdbset persits also in pdbset from ccp4-6.1.3 > > -- > -- > Tim Gruene > Institut fuer anorganische Chemie > Tammannstr. 4 > D-37077 Goettingen > > GPG Key ID = A46BEE1A -- -- Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A signature.asc Description: Digital signature
[ccp4bb] PRACTICAL COURSE ON REACTION KINETICS OF BIOLOGICAL MACROMOLECULES
Dear colleagues, please be informed that online applications are accepted for the following course: 9TH NCCR PRACTICAL COURSE ON REACTION KINETICS OF BIOLOGICAL MACROMOLECULES January 10 - 14, 2011 Zürich, Switzerland http://www.structuralbiology.uzh.ch/nccrcourse2011.asp Course topics include: General introduction to reaction kinetics, reaction kinetics of enzymes, techniques for measuring fast reactions on the millisecond time scale, spectroscopic methods for following reactions between proteins and ligands and conformational transitions in proteins Fluorescence anisotropy measurements, FRET-measurements, single-molecule fluorescence spectroscopy, kinetics of protein folding, evaluation of kinetic data, topical examples from ongoing research projects (invited guest speakers). The main focus of the course will be on practical work in small learning groups. The course is primarily directed to PhD students and postdocs working or planning to work on the reaction mechanism of proteins in vitro. Selection will be based on scientific CV, a questionnaire and recommendation letter. Participants will be invited to suggest their own research protein for course experiments. Interested candidates are encouraged to apply online on http://www.structuralbiology.uzh.ch/nccrcourse2011_application.asp. Application deadline will be October 18, 2010. We will be able to accept 20 participants to this course. Please do not hesitate to contact us if you need any further information. Best regards, Patrick Sticher -- _ Visit the NCCR on the Internet www.structuralbiology.uzh.ch Dr. Patrick Sticher Moser NCCR Scientific Officer Institute of Biochemistry University of Zürich Winterthurerstrasse 190 CH - 8057 Zürich Phone +41 / (0)44 / 635 54 84 Fax +41 / (0)44 / 635 59 08 Mailstic...@bioc.uzh.ch
