Re: [ccp4bb] CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15)
Dear All, thanks for your replies. I tend to believe that it could be ATP some how copurified with the protein. Refinement with ATP, ADP+acetate, did not vary too much in the B factors. The main concern is, only one xtal was observed at this state ( mimicked like a ATP bound state and changed conformation and space group). All other xtals (around 10 xtals were measured) from this plate did not have this change.So it was little tough to understand that only this xtal has ATP instead of ADP. We were not able to reproduce xtals of similar space group with similar setup (with ADP). After tremendous trials we could grow one xtal with ATP which has the P212121 space group and all the observed changes. Is any one observed ATP mimicking by compounds other than aluminium/berillyium Fluoride? Thanks Saravanan Dear All, Sorry for the little bit off topic. Is there a possibility for covalent bond formation between beta phosphate of ADP and acetate molecule both are coordinated by divalent metal ions? I am working on a Kinase structure which was crystallized with ADP and in presence of 1M sodium acetate. We observed additional density around ADP that fits perfectly like a gamma phosphate , mimicking like ATP bound state, surrounded and coordinated by two metal ions(resolution is 1.4A). There is a change in space group (from I212121 to P212121 ) and further important conformation changes are observed around ATP binding pocket and distant region. This is the only xtal we obtained in this space group, and all other xtals(measured 10 xtals) from the same plate belong to I212121. Thanks your help and time! Saravanan Replies: Hello Saravanan At 1.4 Angstrom resolution wouldn't that suggest that you've somehow got ATP in there ? I don't think I understand the other option - were you proposing a ADP-O-C(O)2 arrangement to explain the density ? Surely that has a rather different shape, considerably different scattering power at the center of the terminal group (C vs P) and probably different X-O bond lengths. All of these should show in the density maps at 1.4 Å, although the bond length issue could be quite subtle. Phil Jeffrey Princeton Perhaps it is ATP? Mark J van Raaij Remember that 2xADP can disproportionate into ATP + AMP Hi Saravanan It sounds to me like you have ATP there, even though you added ADP. Is your kinase an inactive mutant? I know kinases can bring nucleotides along for the ride during purification. Regards Christine -- Dr. Saravanan Panneerselvam PhD P11 Beamline Scientist HASYLAB, DESY Building 25F, Room 360 Notkestrasse, 85 22607, Hamburg GERMANY Phone : 0049 40 8998 2692 Mobile: 0049 40 8998 92692 saravananpann...@gmail.com saravanan.panneersel...@desy.de
Re: [ccp4bb] CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15)
Excellent point. I've heard apocryphal stories of someone obtaining beautiful salt crystals (magnesium ammonium phosphate) by trying to crystallize an ATPase with ADP and Mg++ using (NH4)2SO4 as a precipitant. Wait long enough, and the ADP becomes ATP plus AMP, the ATPase takes the 3rd phosphate off, and then the salt crystals grow. Yours sounds like a much less depressing "accident"! From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Phil Evans [p...@mrc-lmb.cam.ac.uk] Sent: Thursday, January 19, 2017 4:37 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15) Remember that 2xADP can disproportionate into ATP + AMP > On 19 Jan 2017, at 20:54, Panneerselvam, Saravanan > <saravanan.panneersel...@desy.de> wrote: > > Dear All, > Sorry for the little bit off topic. Is there a possibility for covalent > bond formation between beta phosphate of ADP and acetate molecule both > are coordinated by divalent metal ions? I am working on a Kinase > structure which was crystallized with ADP and in presence of 1M sodium > acetate. We observed additional density around ADP that fits perfectly > like a gamma phosphate , mimicking like ATP bound state, surrounded and > coordinated by two metal ions(resolution is 1.4A). There is a change in > space group (from I212121 to P212121 ) and further important > conformation changes are observed around ATP binding pocket and distant > region. This is the only xtal we obtained in this space group, and all > other xtals(measured 10 xtals) from the same plate belong to I212121. > > > > Thanks your help and time! > > Saravanan > - Original Message - > From: "CCP4BB automatic digest system" <lists...@jiscmail.ac.uk> > To: CCP4BB@JISCMAIL.AC.UK > Sent: Sunday, 15 January, 2017 01:01:37 > Subject: CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15) > > There are 2 messages totaling 45 lines in this issue. > > Topics of the day: > > 1. Phenix (2) > > -- > > Date:Sat, 14 Jan 2017 20:58:09 + > From:D Bonsor <dbon...@ihv.umaryland.edu> > Subject: Phenix > > Is the phenix website down? Anyone know when it will be back up? > > -- > > Date:Sat, 14 Jan 2017 14:45:06 -0800 > From:Pavel Afonine <pafon...@gmail.com> > Subject: Re: Phenix > > See notice on Phenix mailing list that answers your question. > > On Sat, Jan 14, 2017 at 12:58 PM, D Bonsor <dbon...@ihv.umaryland.edu> > wrote: > >> Is the phenix website down? Anyone know when it will be back up? >> > > -- > > End of CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15) >
Re: [ccp4bb] CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15)
Remember that 2xADP can disproportionate into ATP + AMP > On 19 Jan 2017, at 20:54, Panneerselvam, Saravanan > <saravanan.panneersel...@desy.de> wrote: > > Dear All, > Sorry for the little bit off topic. Is there a possibility for covalent > bond formation between beta phosphate of ADP and acetate molecule both > are coordinated by divalent metal ions? I am working on a Kinase > structure which was crystallized with ADP and in presence of 1M sodium > acetate. We observed additional density around ADP that fits perfectly > like a gamma phosphate , mimicking like ATP bound state, surrounded and > coordinated by two metal ions(resolution is 1.4A). There is a change in > space group (from I212121 to P212121 ) and further important > conformation changes are observed around ATP binding pocket and distant > region. This is the only xtal we obtained in this space group, and all > other xtals(measured 10 xtals) from the same plate belong to I212121. > > > > Thanks your help and time! > > Saravanan > - Original Message - > From: "CCP4BB automatic digest system" <lists...@jiscmail.ac.uk> > To: CCP4BB@JISCMAIL.AC.UK > Sent: Sunday, 15 January, 2017 01:01:37 > Subject: CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15) > > There are 2 messages totaling 45 lines in this issue. > > Topics of the day: > > 1. Phenix (2) > > -- > > Date:Sat, 14 Jan 2017 20:58:09 + > From:D Bonsor <dbon...@ihv.umaryland.edu> > Subject: Phenix > > Is the phenix website down? Anyone know when it will be back up? > > -- > > Date:Sat, 14 Jan 2017 14:45:06 -0800 > From:Pavel Afonine <pafon...@gmail.com> > Subject: Re: Phenix > > See notice on Phenix mailing list that answers your question. > > On Sat, Jan 14, 2017 at 12:58 PM, D Bonsor <dbon...@ihv.umaryland.edu> > wrote: > >> Is the phenix website down? Anyone know when it will be back up? >> > > -- > > End of CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15) >
Re: [ccp4bb] CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15)
On 1/19/17 3:54 PM, Panneerselvam, Saravanan wrote: We observed additional density around ADP that fits perfectly like a gamma phosphate Hello Saravanan At 1.4 Angstrom resolution wouldn't that suggest that you've somehow got ATP in there ? I don't think I understand the other option - were you proposing a ADP-O-C(O)2 arrangement to explain the density ? Surely that has a rather different shape, considerably different scattering power at the center of the terminal group (C vs P) and probably different X-O bond lengths. All of these should show in the density maps at 1.4 Å, although the bond length issue could be quite subtle. Phil Jeffrey Princeton mimicking like ATP bound state, surrounded and coordinated by two metal ions(resolution is 1.4A). There is a change in space group (from I212121 to P212121 ) and further important conformation changes are observed around ATP binding pocket and distant region. This is the only xtal we obtained in this space group, and all other xtals(measured 10 xtals) from the same plate belong to I212121. Thanks your help and time! Saravanan
Re: [ccp4bb] CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15)
Dear All, Sorry for the little bit off topic. Is there a possibility for covalent bond formation between beta phosphate of ADP and acetate molecule both are coordinated by divalent metal ions? I am working on a Kinase structure which was crystallized with ADP and in presence of 1M sodium acetate. We observed additional density around ADP that fits perfectly like a gamma phosphate , mimicking like ATP bound state, surrounded and coordinated by two metal ions(resolution is 1.4A). There is a change in space group (from I212121 to P212121 ) and further important conformation changes are observed around ATP binding pocket and distant region. This is the only xtal we obtained in this space group, and all other xtals(measured 10 xtals) from the same plate belong to I212121. Thanks your help and time! Saravanan - Original Message - From: "CCP4BB automatic digest system" <lists...@jiscmail.ac.uk> To: CCP4BB@JISCMAIL.AC.UK Sent: Sunday, 15 January, 2017 01:01:37 Subject: CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15) There are 2 messages totaling 45 lines in this issue. Topics of the day: 1. Phenix (2) -- Date:Sat, 14 Jan 2017 20:58:09 + From:D Bonsor <dbon...@ihv.umaryland.edu> Subject: Phenix Is the phenix website down? Anyone know when it will be back up? -- Date:Sat, 14 Jan 2017 14:45:06 -0800 From:Pavel Afonine <pafon...@gmail.com> Subject: Re: Phenix See notice on Phenix mailing list that answers your question. On Sat, Jan 14, 2017 at 12:58 PM, D Bonsor <dbon...@ihv.umaryland.edu> wrote: > Is the phenix website down? Anyone know when it will be back up? > -------------- End of CCP4BB Digest - 13 Jan 2017 to 14 Jan 2017 (#2017-15)
