Re: [ccp4bb] On pKa of Aspartic acid
Roger Rowlett wrote:
No. Kw = [H3O+][OH-] = 1 x 10^-14 at 25 deg C.
So at pH 7.0, you have 10^-7 M each at equilibrium no matter how you slice it
or whatever
else is in solution. If equilibrium [H3O+] goes up [OH-] goes down
commensurately.
The pKa of water as an acid is based on Kw and water's effective
concentration of 55 M
in pure water. This pKa is used to compare the instrinsic acidity of water to
other weak
acids. Water is an exceptionally weak acid or base.
And note that Kw, like other physical constants, depends on temperature,
ionic strength, etc. Therefore neutrality, defined as the concentration
where H+ = OH- , which is half of pKw*, is not exactly 7.0, but varies.
A lot of students come out of first-year chemistry with the idea that
pH 7. is by definition neutral.
(*That is using the old Kw definition where {H2O} is taken as 1)
The pK at 14 (or 14+log(55)) is for H2O - OH-, H+
i.e. pH where H2O = OH-
The pK at 0 (or -log 55)) is for H3O+ - H2O, H+
i.e. pH ph when H3O+ = H2O
But isn't H+ = H30? then when H3O+ = H2O, [H3O] = 55/2,
pH would be -log (55/2)
pH is log of activity of water, or whatever the glass electrode measure.
Or- assuming [H2O] always unity, when H2O = H3O+, H3O+ = 1, pH=0
assuming [H2O] always 55, extrapolate to where H3O+ = H2O while keeping H20=55,
then pH would be -log 55
Re: [ccp4bb] On pKa of Aspartic acid
Hi Deepak, Assuming that you have done the necessary things to measure the pKr of that particular Asp, I would say that the increase is advantageous for your enzyme. Enzyme catalysis often involves very subtle changes on the ionization state of the active site. But you need to be very careful in proposing a catalysis mechanism. b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? Are you sure that the water donates a proton? Was your resolution high enough to observe it? How did you measure that pKr, by the way? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? I would say that the dominant fraction of Asp is deprotonated. But as you can see in the papers below, the pKr of Asp can vary from 0.5 to 9.2 in folded proteins. d) Have similar increase in pKa values observed for aspartic acids before? there are some papers by Nick Pace's group: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2708032/?tool=pubmed http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2679426/?tool=pubmed http://www.sciencedirect.com/science/article/pii/S002228360600934X Cheers, Clement -- On 2/7/12 8:48 PM, Deepak Oswal wrote: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Hi Deepak: I think it is common for the residues which participate catalysis to have a Pka deviated from the reality pKa value especially for acid/base catalysis (acid base titration assay can help you to figure out the way of catalysis). Usually the pKa values of these kind of critical residues are affected by their local environment and this character is related to the enzyme's working mechanism. I am sorry that I am not professional in enzyme, I cannot answer your questions for each questions. Yu Xiaodi Date: Tue, 7 Feb 2012 19:48:26 +0800 From: deepos...@gmail.com Subject: [ccp4bb] On pKa of Aspartic acid To: CCP4BB@JISCMAIL.AC.UK Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Residue pKa values in proteins are strongly affected by the local environment and can deviate far from the "norm". Of course, the higher the pKa of the residue, the stronger general base it will be. There is a significant thermodynamic/kinetic advantage in matching the pKa of a general base to the pKa of the proton that must be removed for catalysis. Aspartate frequently plays a role as a general base for assisting water in nucleophilic attack. I think it is unusual for water to act as a general acid (the pKa of water is 14-15, depending on how you calculate it), but if that is the case and there is compelling evidence that water is the proton donor in your reaction, it would be necessary for aspartic acid to be in its protonated form in order to assist a water-mediated protonation event. Raising the pKa of aspartic acid would allow a larger fraction of it to be in its protonated state at physiologically relevant pH values, although it would reduce the intrinsic effectiveness of Asp as a general acid. There should be a significant thermodynamic and kinetic advantage in having Asp participate directly in a general acid catalyzed reaction, rather than through a water molecule. Cheers, ___ Roger S. Rowlett Gordon Dorothy Kline Professor Department of Chemistry Colgate University 13 Oak Drive Hamilton, NY 13346 tel: (315)-228-7245 ofc: (315)-228-7395 fax: (315)-228-7935 email: rrowl...@colgate.edu On 2/7/2012 10:00 AM, Xiaodi Yu wrote: Hi Deepak: I think it is common for the residues which participate catalysis to have a Pka deviated from the reality pKa value especially for acid/base catalysis (acid base titration assay can help you to figure out the way of catalysis). Usually the pKa values of these kind of critical residues are affected by their local environment and this character is related to the enzyme's working mechanism. I am sorry that I am not professional in enzyme, I cannot answer your questions for each questions. Yu Xiaodi Date: Tue, 7 Feb 2012 19:48:26 +0800 From: deepos...@gmail.com Subject: [ccp4bb] On pKa of Aspartic acid To: CCP4BB@JISCMAIL.AC.UK Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
As we know, the pKa of water is 15.7. Under pH 7.0, its protonation should be 50/50. In this case, we may need to consider water in two formats: H2O vs. H3O+ When we say water as acid, it usually stands for H3O+ in chemistry. In chemical equation, H+ represents H3O+. In enzyme catalysis, water as a general acid sounds reasonable under pH 7.0. In some famous paper, water has been concluded as the general base (pKa 15.7) to deprotonate an alpha hydrogen (pKa ~ 22) or a hydrogen from a sp3 hybridized carbon (pKa ~36). This logic may need to be reconsidered. . Recently, I have read papers for pKa perturbation. I am also interested in the general base of Asp and Glu in enzyme catalysis. I will be very happy to read your paper in the future. Regards, Kevin Jin On Tue, Feb 7, 2012 at 3:48 AM, Deepak Oswal deepos...@gmail.com wrote: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Check this review, for instance: Pace, C. et al. Protein Ionizable Groups: pK values and Their Contribution to Protein Stability and Solubility. J. Biol Chem. 284, 13285-13289 (May 15, 2009) Thierry From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Xiaodi Yu Sent: Tuesday, February 07, 2012 10:00 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] On pKa of Aspartic acid Hi Deepak: I think it is common for the residues which participate catalysis to have a Pka deviated from the reality pKa value especially for acid/base catalysis (acid base titration assay can help you to figure out the way of catalysis). Usually the pKa values of these kind of critical residues are affected by their local environment and this character is related to the enzyme's working mechanism. I am sorry that I am not professional in enzyme, I cannot answer your questions for each questions. Yu Xiaodi Date: Tue, 7 Feb 2012 19:48:26 +0800 From: deepos...@gmail.com Subject: [ccp4bb] On pKa of Aspartic acid To: CCP4BB@JISCMAIL.AC.UK Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids' ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn't that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal Notice: This e-mail message, together with any attachments, contains information of Merck Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates Direct contact information for affiliates is available at http://www.merck.com/contact/contacts.html) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system.
Re: [ccp4bb] On pKa of Aspartic acid
Oops, you meant catalytic residue. Check the following: Harris TK Turner GJ Structural basis of pertubed pKa values of catalytic groups in enzyme active sites IUBMB life, 53 85-98 (Feb 2002) Thierry From: Fischmann, Thierry Sent: Tuesday, February 07, 2012 10:59 AM To: 'Xiaodi Yu'; CCP4BB@JISCMAIL.AC.UK Subject: RE: [ccp4bb] On pKa of Aspartic acid Check this review, for instance: Pace, C. et al. Protein Ionizable Groups: pK values and Their Contribution to Protein Stability and Solubility. J. Biol Chem. 284, 13285-13289 (May 15, 2009) Thierry From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Xiaodi Yu Sent: Tuesday, February 07, 2012 10:00 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] On pKa of Aspartic acid Hi Deepak: I think it is common for the residues which participate catalysis to have a Pka deviated from the reality pKa value especially for acid/base catalysis (acid base titration assay can help you to figure out the way of catalysis). Usually the pKa values of these kind of critical residues are affected by their local environment and this character is related to the enzyme's working mechanism. I am sorry that I am not professional in enzyme, I cannot answer your questions for each questions. Yu Xiaodi Date: Tue, 7 Feb 2012 19:48:26 +0800 From: deepos...@gmail.com Subject: [ccp4bb] On pKa of Aspartic acid To: CCP4BB@JISCMAIL.AC.UK Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids' ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn't that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal Notice: This e-mail message, together with any attachments, contains information of Merck Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates Direct contact information for affiliates is available at http://www.merck.com/contact/contacts.html) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system.
Re: [ccp4bb] On pKa of Aspartic acid
Hi, you may also look into the papers of John A. Gerlt, who did a lot on protonabtraction reactions and the theory behind this. Esspecially the pKa disturbance and the match to the pkA of the substrate of the reaction. Best Wishes Christian Am Dienstag 07 Februar 2012 12:48:26 schrieb Deepak Oswal: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Is the water molecule in question coordinated to any other group(s)? -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Kevin Jin Sent: Tuesday, February 07, 2012 10:22 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] On pKa of Aspartic acid As we know, the pKa of water is 15.7. Under pH 7.0, its protonation should be 50/50. In this case, we may need to consider water in two formats: H2O vs. H3O+ When we say water as acid, it usually stands for H3O+ in chemistry. In chemical equation, H+ represents H3O+. In enzyme catalysis, water as a general acid sounds reasonable under pH 7.0. In some famous paper, water has been concluded as the general base (pKa 15.7) to deprotonate an alpha hydrogen (pKa ~ 22) or a hydrogen from a sp3 hybridized carbon (pKa ~36). This logic may need to be reconsidered. . Recently, I have read papers for pKa perturbation. I am also interested in the general base of Asp and Glu in enzyme catalysis. I will be very happy to read your paper in the future. Regards, Kevin Jin On Tue, Feb 7, 2012 at 3:48 AM, Deepak Oswal deepos...@gmail.com wrote: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids' ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn't that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Maybe you would also be interested in http://www.jinkai.org/AAD_history.html Regards, Kevin On Tue, Feb 7, 2012 at 8:52 AM, Christian Roth christian.r...@bbz.uni-leipzig.de wrote: Hi, you may also look into the papers of John A. Gerlt, who did a lot on protonabtraction reactions and the theory behind this. Esspecially the pKa disturbance and the match to the pkA of the substrate of the reaction. Best Wishes Christian Am Dienstag 07 Februar 2012 12:48:26 schrieb Deepak Oswal: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Hi Deepak, With regards observed pKa shifts, Prof. Ondrechen from Northeastern University has had a long interest in this field. http://www.northeastern.edu/org/wp/ Under the computational tools that she has developed a program called THEMATICS that allows you to predict the pka of titratable amino acids and she has been able to predict shifts. Though the server seems to be down at this point, here is the reference: Y. Wei, J. Ko, L.F. Murga, and M.J. Ondrechen, BMC Bioinformatics 8:119, (2007) From the commercial side, Dr. Spassov from Accelrys has also been working on tools that predict protein ionization. In his work, he has also been able to predict significant pka shifts for functionally relevant residues. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2578799/ Cheers, Francisco From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Deepak Oswal Sent: Tuesday, February 07, 2012 3:48 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] On pKa of Aspartic acid Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids' ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn't that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Hi Kevin, Hate to point this out, but under pH 7.0, the protonation state of water is not 50:50, and it is not a good acid. The H30+ concentration of pure water is 10^-7 Molar. In pure water (assuming 55.5 M) only 1:555,000,000 water molecules is in the protonated, charged state (H3O+). This is why when an enzyme uses water in its mechanism as a nucleophile, base, or acid, there is usually an acid/base catalyst or metal that protonates or deprotonates the water to 'activate it'. Best regards, Z *** Zachary A. Wood, Ph.D. Assistant Professor Department of Biochemistry Molecular Biology University of Georgia Life Sciences Building, Rm A426B 120 Green Street Athens, GA 30602-7229 Office: 706-583-0304 Lab:706-583-0303 FAX: 706-542-1738 *** On Feb 7, 2012, at 11:22 AM, Kevin Jin wrote: As we know, the pKa of water is 15.7. Under pH 7.0, its protonation should be 50/50. In this case, we may need to consider water in two formats: H2O vs. H3O+ When we say water as acid, it usually stands for H3O+ in chemistry. In chemical equation, H+ represents H3O+. In enzyme catalysis, water as a general acid sounds reasonable under pH 7.0. In some famous paper, water has been concluded as the general base (pKa 15.7) to deprotonate an alpha hydrogen (pKa ~ 22) or a hydrogen from a sp3 hybridized carbon (pKa ~36). This logic may need to be reconsidered. . Recently, I have read papers for pKa perturbation. I am also interested in the general base of Asp and Glu in enzyme catalysis. I will be very happy to read your paper in the future. Regards, Kevin Jin On Tue, Feb 7, 2012 at 3:48 AM, Deepak Oswal deepos...@gmail.com wrote: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Oops, It should be: [H3O+]/[OH-]= 50/50 Kw = [H3O+][OH-], pH = pKa +log ([OH-]/[H2O]) H3O+ concentration of pure water is 10^-7 mol/L total H+ = 55.5M * 10^-7 = 5.55* 10^-6 mole. Is this right? Regards, Kevin On Tue, Feb 7, 2012 at 12:13 PM, Zachary Wood z...@bmb.uga.edu wrote: Hi Kevin, Hate to point this out, but under pH 7.0, the protonation state of water is not 50:50, and it is not a good acid. The H30+ concentration of pure water is 10^-7 Molar. In pure water (assuming 55.5 M) only 1:555,000,000 water molecules is in the protonated, charged state (H3O+). This is why when an enzyme uses water in its mechanism as a nucleophile, base, or acid, there is usually an acid/base catalyst or metal that protonates or deprotonates the water to 'activate it'. Best regards, Z *** Zachary A. Wood, Ph.D. Assistant Professor Department of Biochemistry Molecular Biology University of Georgia Life Sciences Building, Rm A426B 120 Green Street Athens, GA 30602-7229 Office: 706-583-0304 Lab: 706-583-0303 FAX: 706-542-1738 *** On Feb 7, 2012, at 11:22 AM, Kevin Jin wrote: As we know, the pKa of water is 15.7. Under pH 7.0, its protonation should be 50/50. In this case, we may need to consider water in two formats: H2O vs. H3O+ When we say water as acid, it usually stands for H3O+ in chemistry. In chemical equation, H+ represents H3O+. In enzyme catalysis, water as a general acid sounds reasonable under pH 7.0. In some famous paper, water has been concluded as the general base (pKa 15.7) to deprotonate an alpha hydrogen (pKa ~ 22) or a hydrogen from a sp3 hybridized carbon (pKa ~36). This logic may need to be reconsidered. . Recently, I have read papers for pKa perturbation. I am also interested in the general base of Asp and Glu in enzyme catalysis. I will be very happy to read your paper in the future. Regards, Kevin Jin On Tue, Feb 7, 2012 at 3:48 AM, Deepak Oswal deepos...@gmail.com wrote: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
No. Kw = [H3O+][OH-] = 1 x 10^-14 at 25 deg C. So at pH 7.0, you have 10^-7 M each at equilibrium no matter how you slice it or whatever else is in solution. If equilibrium [H3O+] goes up [OH-] goes down commensurately. The "pKa" of water as an acid is based on Kw and water's effective concentration of 55 M in pure water. This "pKa" is used to compare the instrinsic acidity of water to other weak acids. Water is an exceptionally weak acid or base. ___ Roger S. Rowlett Gordon Dorothy Kline Professor Department of Chemistry Colgate University 13 Oak Drive Hamilton, NY 13346 tel: (315)-228-7245 ofc: (315)-228-7395 fax: (315)-228-7935 email: rrowl...@colgate.edu On 2/7/2012 3:42 PM, Kevin Jin wrote: Oops, It should be: [H3O+]/[OH-]= 50/50 Kw = [H3O+][OH-], pH = pKa +log ([OH-]/[H2O]) H3O+ concentration of pure water is 10^-7 mol/L total H+ = 55.5M * 10^-7 = 5.55* 10^-6 mole. Is this right? Regards, Kevin On Tue, Feb 7, 2012 at 12:13 PM, Zachary Wood z...@bmb.uga.edu wrote: Hi Kevin, Hate to point this out, but under pH 7.0, the protonation state of water is not 50:50, and it is not a good acid. The H30+ concentration of pure water is 10^-7 Molar. In pure water (assuming 55.5 M) only 1:555,000,000 water molecules is in the protonated, charged state (H3O+). This is why when an enzyme uses water in its mechanism as a nucleophile, base, or acid, there is usually an acid/base catalyst or metal that protonates or deprotonates the water to 'activate it'. Best regards, Z *** Zachary A. Wood, Ph.D. Assistant Professor Department of Biochemistry Molecular Biology University of Georgia Life Sciences Building, Rm A426B 120 Green Street Athens, GA 30602-7229 Office: 706-583-0304 Lab: 706-583-0303 FAX: 706-542-1738 *** On Feb 7, 2012, at 11:22 AM, Kevin Jin wrote: As we know, the pKa of water is 15.7. Under pH 7.0, its protonation should be 50/50. In this case, we may need to consider water in two formats: H2O vs. H3O+ When we say water as acid, it usually stands for H3O+ in chemistry. In chemical equation, H+ represents H3O+. In enzyme catalysis, water as a general acid sounds reasonable under pH 7.0. In some famous paper, water has been concluded as the general base (pKa 15.7) to deprotonate an alpha hydrogen (pKa ~ 22) or a hydrogen from a sp3 hybridized carbon (pKa ~36). This logic may need to be reconsidered. . Recently, I have read papers for pKa perturbation. I am also interested in the general base of Asp and Glu in enzyme catalysis. I will be very happy to read your paper in the future. Regards, Kevin Jin On Tue, Feb 7, 2012 at 3:48 AM, Deepak Oswal deepos...@gmail.com wrote: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal
Re: [ccp4bb] On pKa of Aspartic acid
Dear all, for further discussion I believe that using the 0-14 pH scale assumes water activity of pure water, something that is surely not matched in the surface or pocket of a protein, so keeping this in mind I always prefer to speak about apparent pKa of a group if talking about a non solvent-exposed group. Horacio 2012/2/7 Roger Rowlett rrowl...@colgate.edu No. Kw = [H3O+][OH-] = 1 x 10^-14 at 25 deg C. So at pH 7.0, you have 10^-7 M each at equilibrium no matter how you slice it or whatever else is in solution. If equilibrium [H3O+] goes up [OH-] goes down commensurately. The pKa of water as an acid is based on Kw and water's effective concentration of 55 M in pure water. This pKa is used to compare the instrinsic acidity of water to other weak acids. Water is an exceptionally weak acid or base. ___ Roger S. Rowlett Gordon Dorothy Kline Professor Department of Chemistry Colgate University 13 Oak Drive Hamilton, NY 13346 tel: (315)-228-7245 ofc: (315)-228-7395 fax: (315)-228-7935 email: rrowl...@colgate.edu On 2/7/2012 3:42 PM, Kevin Jin wrote: Oops, It should be: [H3O+]/[OH-]= 50/50 Kw = [H3O+][OH-], pH = pKa +log ([OH-]/[H2O]) H3O+ concentration of pure water is 10^-7 mol/L total H+ = 55.5M * 10^-7 = 5.55* 10^-6 mole. Is this right? Regards, Kevin On Tue, Feb 7, 2012 at 12:13 PM, Zachary Wood z...@bmb.uga.edu z...@bmb.uga.edu wrote: Hi Kevin, Hate to point this out, but under pH 7.0, the protonation state of water is not 50:50, and it is not a good acid. The H30+ concentration of pure water is 10^-7 Molar. In pure water (assuming 55.5 M) only 1:555,000,000 water molecules is in the protonated, charged state (H3O+). This is why when an enzyme uses water in its mechanism as a nucleophile, base, or acid, there is usually an acid/base catalyst or metal that protonates or deprotonates the water to 'activate it'. Best regards, Z *** Zachary A. Wood, Ph.D. Assistant Professor Department of Biochemistry Molecular Biology University of Georgia Life Sciences Building, Rm A426B 120 Green Street Athens, GA 30602-7229 Office: 706-583-0304 Lab:706-583-0303 FAX: 706-542-1738 *** On Feb 7, 2012, at 11:22 AM, Kevin Jin wrote: As we know, the pKa of water is 15.7. Under pH 7.0, its protonation should be 50/50. In this case, we may need to consider water in two formats: H2O vs. H3O+ When we say water as acid, it usually stands for H3O+ in chemistry. In chemical equation, H+ represents H3O+. In enzyme catalysis, water as a general acid sounds reasonable under pH 7.0. In some famous paper, water has been concluded as the general base (pKa 15.7) to deprotonate an alpha hydrogen (pKa ~ 22) or a hydrogen from a sp3 hybridized carbon (pKa ~36). This logic may need to be reconsidered. . Recently, I have read papers for pKa perturbation. I am also interested in the general base of Asp and Glu in enzyme catalysis. I will be very happy to read your paper in the future. Regards, Kevin Jin On Tue, Feb 7, 2012 at 3:48 AM, Deepak Oswal deepos...@gmail.com deepos...@gmail.com wrote: Dear colleagues, We have solved the crystal structure of a human enzyme. The pKa of a catalytically critical aspartic acid has increased to 6.44. It is hydrogen bonded (2.8 Angstroms) to a water molecule that is supposed to donate a proton during the catalysis. Can anybody help me a) interpret the significance of this increase in pKa of the aspartic acid from 3.8 to 6.44 in context with the catalysis? Is this advantageous or detrimental? b) How is pKa related to an amino acids’ ability to force a water molecule to donate a proton? c) At pH 7.4, the aspartic acid would be de-protonated irrespective of whether the pKa is 3.8 or 6.44; isn’t that true? d) Have similar increase in pKa values observed for aspartic acids before? I would be grateful if anybody could explain or comment on the above queries. Deepak Oswal -- Horacio Botti PhD MD Protein Crystallography Unit Institut Pasteur of Montevideo
