1zvk
From Proteopedia
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|PDB= 1zvk |SIZE=350|CAPTION= <scene name='initialview01'>1zvk</scene>, resolution 2.04Å | |PDB= 1zvk |SIZE=350|CAPTION= <scene name='initialview01'>1zvk</scene>, resolution 2.04Å | ||
|SITE= | |SITE= | ||
| - | |LIGAND= <scene name='pdbligand=CA:CALCIUM ION'>CA</scene> | + | |LIGAND= <scene name='pdbligand=CA:CALCIUM+ION'>CA</scene> |
|ACTIVITY= | |ACTIVITY= | ||
|GENE= | |GENE= | ||
| + | |DOMAIN= | ||
| + | |RELATEDENTRY=[[1s1o|1S1O]], [[1s1u|1S1U]], [[1sn7|1SN7]], [[1zvk|1ZVK]] | ||
| + | |RESOURCES=<span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=1zvk FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1zvk OCA], [http://www.ebi.ac.uk/pdbsum/1zvk PDBsum], [http://www.rcsb.org/pdb/explore.do?structureId=1zvk RCSB]</span> | ||
}} | }} | ||
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[[Category: Nakayama, T.]] | [[Category: Nakayama, T.]] | ||
[[Category: Wlodawer, A.]] | [[Category: Wlodawer, A.]] | ||
| - | + | [[Category: d164n,e78h]] | |
| - | [[Category: d164n | + | |
| - | + | ||
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Mar 31 01:42:13 2008'' |
Revision as of 22:42, 30 March 2008
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| , resolution 2.04Å | |||||||
|---|---|---|---|---|---|---|---|
| Ligands: | |||||||
| Related: | 1S1O, 1S1U, 1SN7, 1ZVK
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| Resources: | FirstGlance, OCA, PDBsum, RCSB | ||||||
| Coordinates: | save as pdb, mmCIF, xml | ||||||
Structure of Double mutant, D164N, E78H of Kumamolisin-As
Overview
Kumamolisin-As is an acid collagenase with a subtilisin-like fold. Its active site contains a unique catalytic triad, Ser278-Glu78-Asp82, and a putative transition-state stabilizing residue, Asp164. In this study, the mutants D164N and E78H/D164N were engineered in order to replace parts of the catalytic machinery of kumamolisin-As with the residues found in the equivalent positions in subtilisin. Unlike the wild-type and D164N proenzymes, which undergo instantaneous processing to produce their 37-kDa mature forms, the expressed E78H/D164N proenzyme exists as an equilibrated mixture of the nicked and intact forms of the precursor. X-ray crystallographic structures of the mature forms of the two mutants showed that, in each of them, the catalytic Ser278 makes direct hydrogen bonds with the side chain of Asn164. In addition, His78 of the double mutant is distant from Ser278 and Asp82, and the catalytic triad no longer exists. Consistent with these structural alterations around the active site, these mutants showed only low catalytic activity (relative k(cat) at pH 4.0 1.3% for D164N and 0.0001% for E78H/D164N). pH-dependent kinetic studies showed that the single D164N substitution did not significantly alter the logk(cat) vs. pH and log(k(cat)/Km) vs. pH profiles of the enzyme. In contrast, the double mutation resulted in a dramatic switch of the logk(cat) vs. pH profile to one that was consistent with catalysis by means of the Ser278-His78 dyad and Asn164, which may also account for the observed ligation/cleavage equilibrium of the precursor of E78H/D164N. These results corroborate the mechanistic importance of the glutamate-mediated catalytic triad and oxyanion-stabilizing aspartic acid residue for low-pH peptidase activity of the enzyme.
About this Structure
1ZVK is a Single protein structure of sequence from Alicyclobacillus sendaiensis. Full crystallographic information is available from OCA.
Reference
Processing, catalytic activity and crystal structures of kumamolisin-As with an engineered active site., Okubo A, Li M, Ashida M, Oyama H, Gustchina A, Oda K, Dunn BM, Wlodawer A, Nakayama T, FEBS J. 2006 Jun;273(11):2563-76. PMID:16704427
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