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2h5c
From Proteopedia
(New page: 200px<br /><applet load="2h5c" size="450" color="white" frame="true" align="right" spinBox="true" caption="2h5c, resolution 0.82Å" /> '''0.82A resolution cry...) |
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| - | [[Image:2h5c.gif|left|200px]]<br /><applet load="2h5c" size=" | + | [[Image:2h5c.gif|left|200px]]<br /><applet load="2h5c" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="2h5c, resolution 0.82Å" /> | caption="2h5c, resolution 0.82Å" /> | ||
'''0.82A resolution crystal structure of alpha-lytic protease at pH 5'''<br /> | '''0.82A resolution crystal structure of alpha-lytic protease at pH 5'''<br /> | ||
==Overview== | ==Overview== | ||
| - | To address questions regarding the mechanism of serine protease catalysis, we have solved two X-ray crystal structures of alpha-lytic protease | + | To address questions regarding the mechanism of serine protease catalysis, we have solved two X-ray crystal structures of alpha-lytic protease (alphaLP) that mimic aspects of the transition states: alphaLP at pH 5 (0.82 A resolution) and alphaLP bound to the peptidyl boronic acid inhibitor, MeOSuc-Ala-Ala-Pro-boroVal (0.90 A resolution). Based on these structures, there is no evidence of, or requirement for, histidine-flipping during the acylation step of the reaction. Rather, our data suggests that upon protonation of His57, Ser195 undergoes a conformational change that destabilizes the His57-Ser195 hydrogen bond, preventing the back-reaction. In both structures the His57-Asp102 hydrogen bond in the catalytic triad is a normal ionic hydrogen bond, and not a low-barrier hydrogen bond (LBHB) as previously hypothesized. We propose that the enzyme has evolved a network of relatively short hydrogen bonds that collectively stabilize the transition states. In particular, a short ionic hydrogen bond (SIHB) between His57 Nepsilon2 and the substrate's leaving group may promote forward progression of the TI1-to-acylenzyme reaction. We provide experimental evidence that refutes use of either a short donor-acceptor distance or a downfield 1H chemical shift as sole indicators of a LBHB. |
==About this Structure== | ==About this Structure== | ||
| - | 2H5C is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Lysobacter_enzymogenes Lysobacter enzymogenes] with SO4 and GOL as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Alpha-lytic_endopeptidase Alpha-lytic endopeptidase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.4.21.12 3.4.21.12] Full crystallographic information is available from [http:// | + | 2H5C is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Lysobacter_enzymogenes Lysobacter enzymogenes] with <scene name='pdbligand=SO4:'>SO4</scene> and <scene name='pdbligand=GOL:'>GOL</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Alpha-lytic_endopeptidase Alpha-lytic endopeptidase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.4.21.12 3.4.21.12] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2H5C OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Lysobacter enzymogenes]] | [[Category: Lysobacter enzymogenes]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Agard, D | + | [[Category: Agard, D A.]] |
| - | [[Category: Daugherty, M | + | [[Category: Daugherty, M D.]] |
| - | [[Category: Fuhrmann, C | + | [[Category: Fuhrmann, C N.]] |
[[Category: GOL]] | [[Category: GOL]] | ||
[[Category: SO4]] | [[Category: SO4]] | ||
| Line 28: | Line 28: | ||
[[Category: ultra-high resolution]] | [[Category: ultra-high resolution]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 17:38:29 2008'' |
Revision as of 15:38, 21 February 2008
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0.82A resolution crystal structure of alpha-lytic protease at pH 5
Overview
To address questions regarding the mechanism of serine protease catalysis, we have solved two X-ray crystal structures of alpha-lytic protease (alphaLP) that mimic aspects of the transition states: alphaLP at pH 5 (0.82 A resolution) and alphaLP bound to the peptidyl boronic acid inhibitor, MeOSuc-Ala-Ala-Pro-boroVal (0.90 A resolution). Based on these structures, there is no evidence of, or requirement for, histidine-flipping during the acylation step of the reaction. Rather, our data suggests that upon protonation of His57, Ser195 undergoes a conformational change that destabilizes the His57-Ser195 hydrogen bond, preventing the back-reaction. In both structures the His57-Asp102 hydrogen bond in the catalytic triad is a normal ionic hydrogen bond, and not a low-barrier hydrogen bond (LBHB) as previously hypothesized. We propose that the enzyme has evolved a network of relatively short hydrogen bonds that collectively stabilize the transition states. In particular, a short ionic hydrogen bond (SIHB) between His57 Nepsilon2 and the substrate's leaving group may promote forward progression of the TI1-to-acylenzyme reaction. We provide experimental evidence that refutes use of either a short donor-acceptor distance or a downfield 1H chemical shift as sole indicators of a LBHB.
About this Structure
2H5C is a Single protein structure of sequence from Lysobacter enzymogenes with and as ligands. Active as Alpha-lytic endopeptidase, with EC number 3.4.21.12 Full crystallographic information is available from OCA.
Reference
Subangstrom crystallography reveals that short ionic hydrogen bonds, and not a His-Asp low-barrier hydrogen bond, stabilize the transition state in serine protease catalysis., Fuhrmann CN, Daugherty MD, Agard DA, J Am Chem Soc. 2006 Jul 19;128(28):9086-102. PMID:16834383
Page seeded by OCA on Thu Feb 21 17:38:29 2008
Categories: Alpha-lytic endopeptidase | Lysobacter enzymogenes | Single protein | Agard, D A. | Daugherty, M D. | Fuhrmann, C N. | GOL | SO4 | A-lytic protease | Acylation transition state | Catalysis | Packing distortion | Protein folding | Protein stability | Serine protease | Ultra-high resolution
