From Proteopedia
(Difference between revisions)
proteopedia linkproteopedia link
|
|
| Line 1: |
Line 1: |
| - | [[Image:1dmn.jpg|left|200px]] | + | {{Seed}} |
| | + | [[Image:1dmn.png|left|200px]] |
| | | | |
| | <!-- | | <!-- |
| Line 9: |
Line 10: |
| | {{STRUCTURE_1dmn| PDB=1dmn | SCENE= }} | | {{STRUCTURE_1dmn| PDB=1dmn | SCENE= }} |
| | | | |
| - | '''CRYSTAL STRUCTURE OF MUTANT ENZYME Y32F/Y57F OF KETOSTEROID ISOMERASE FROM PSEUDOMONAS PUTIDA BIOTYPE B'''
| + | ===CRYSTAL STRUCTURE OF MUTANT ENZYME Y32F/Y57F OF KETOSTEROID ISOMERASE FROM PSEUDOMONAS PUTIDA BIOTYPE B=== |
| | | | |
| | | | |
| - | ==Overview==
| + | <!-- |
| - | Delta(5)-3-Ketosteroid isomerase from Pseudomonas putida biotype B is one of the most proficient enzymes catalyzing an allylic isomerization reaction at rates comparable to the diffusion limit. The hydrogen-bond network (Asp99... Wat504...Tyr14...Tyr55...Tyr30) which links the two catalytic residues, Tyr14 and Asp99, to Tyr30, Tyr55, and a water molecule in the highly apolar active site has been characterized in an effort to identify its roles in function and stability. The DeltaG(U)(H2O) determined from equilibrium unfolding experiments reveals that the elimination of the hydroxyl group of Tyr14 or Tyr55 or the replacement of Asp99 with leucine results in a loss of conformational stability of 3.5-4.4 kcal/mol, suggesting that the hydrogen bonds of Tyr14, Tyr55, and Asp99 contribute significantly to stability. While decreasing the stability by about 6.5-7.9 kcal/mol, the Y55F/D99L or Y30F/D99L double mutation also reduced activity significantly, exhibiting a synergistic effect on k(cat) relative to the respective single mutations. These results indicate that the hydrogen-bond network is important for both stability and function. Additionally, they suggest that Tyr14 cannot function efficiently alone without additional support from the hydrogen bonds of Tyr55 and Asp99. The crystal structure of Y55F as determined at 1.9 A resolution shows that Tyr14 OH undergoes an alteration in orientation to form a new hydrogen bond with Tyr30. This observation supports the role of Tyr55 OH in positioning Tyr14 properly to optimize the hydrogen bond between Tyr14 and C3-O of the steroid substrate. No significant structural changes were observed in the crystal structures of Y30F and Y30F/Y55F, which allowed us to estimate approximately the interaction energies mediated by the hydrogen bonds Tyr30...Tyr55 and Tyr14...Tyr55. Taken together, our results demonstrate that the hydrogen-bond network provides the structural support that is needed for the enzyme to maintain the active-site geometry optimized for both function and stability.
| + | The line below this paragraph, {{ABSTRACT_PUBMED_10769113}}, adds the Publication Abstract to the page |
| | + | (as it appears on PubMed at http://www.pubmed.gov), where 10769113 is the PubMed ID number. |
| | + | --> |
| | + | {{ABSTRACT_PUBMED_10769113}} |
| | | | |
| | ==About this Structure== | | ==About this Structure== |
| Line 31: |
Line 35: |
| | [[Category: Coneshell]] | | [[Category: Coneshell]] |
| | [[Category: Curved b-sheet]] | | [[Category: Curved b-sheet]] |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Fri May 2 14:01:42 2008'' | + | |
| | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Jun 30 23:18:05 2008'' |
Revision as of 20:18, 30 June 2008
Template:STRUCTURE 1dmn
CRYSTAL STRUCTURE OF MUTANT ENZYME Y32F/Y57F OF KETOSTEROID ISOMERASE FROM PSEUDOMONAS PUTIDA BIOTYPE B
Template:ABSTRACT PUBMED 10769113
About this Structure
1DMN is a Single protein structure of sequence from Pseudomonas putida. Full crystallographic information is available from OCA.
Reference
Contribution of the hydrogen-bond network involving a tyrosine triad in the active site to the structure and function of a highly proficient ketosteroid isomerase from Pseudomonas putida biotype B., Kim DH, Jang DS, Nam GH, Choi G, Kim JS, Ha NC, Kim MS, Oh BH, Choi KY, Biochemistry. 2000 Apr 25;39(16):4581-9. PMID:10769113
Page seeded by OCA on Mon Jun 30 23:18:05 2008
