|
|
| Line 1: |
Line 1: |
| [[Image:1bgn.gif|left|200px]] | | {{Seed}} |
| | [[Image:1bgn.png|left|200px]] |
|
| |
|
| <!-- | | <!-- |
| Line 9: |
Line 10: |
| {{STRUCTURE_1bgn| PDB=1bgn | SCENE= }} | | {{STRUCTURE_1bgn| PDB=1bgn | SCENE= }} |
|
| |
|
| '''P-HYDROXYBENZOATE HYDROXYLASE (PHBH) MUTANT WITH CYS 116 REPLACED BY SER (C116S) AND ARG 269 REPLACED BY THR (R269T), IN COMPLEX WITH FAD AND 4-HYDROXYBENZOIC ACID'''
| | ===P-HYDROXYBENZOATE HYDROXYLASE (PHBH) MUTANT WITH CYS 116 REPLACED BY SER (C116S) AND ARG 269 REPLACED BY THR (R269T), IN COMPLEX WITH FAD AND 4-HYDROXYBENZOIC ACID=== |
|
| |
|
|
| |
|
| ==Overview==
| | <!-- |
| The conserved residues His-162 and Arg-269 of the flavoprotein p-hydroxybenzoate hydroxylase (EC 1.14.13.2) are located at the entrance of the interdomain cleft that leads toward the active site. To study their putative role in NADPH binding, His-162 and Arg-269 were selectively changed by site-specific mutagenesis. The catalytic properties of H162R, H162Y, and R269K were similar to the wild-type enzyme. However, less conservative His-162 and Arg-269 replacements strongly impaired NADPH binding without affecting the conformation of the flavin ring and the efficiency of substrate hydroxylation. The crystal structures of H162R and R269T in complex with 4-hydroxybenzoate were solved at 3.0 and 2.0 A resolution, respectively. Both structures are virtually indistinguishable from the wild-type enzyme-substrate complex except for the substituted side chains. In contrast to wild-type p-hydroxybenzoate hydroxylase, H162R is not inactivated by diethyl pyrocarbonate. NADPH protects wild-type p-hydroxybenzoate hydroxylase from diethylpyrocarbonate inactivation, suggesting that His-162 is involved in NADPH binding. Based on these results and GRID calculations we propose that the side chains of His-162 and Arg-269 interact with the pyrophosphate moiety of NADPH. An interdomain binding mode for NADPH is proposed which takes a novel sequence motif (Eppink, M. H. M., Schreuder, H. A., and van Berkel, W. J. H. (1997) Protein Sci. 6, 2454-2458) into account.
| | The line below this paragraph, {{ABSTRACT_PUBMED_9694855}}, adds the Publication Abstract to the page |
| | (as it appears on PubMed at http://www.pubmed.gov), where 9694855 is the PubMed ID number. |
| | --> |
| | {{ABSTRACT_PUBMED_9694855}} |
|
| |
|
| ==About this Structure== | | ==About this Structure== |
| Line 27: |
Line 31: |
| [[Category: Schreuder, H A.]] | | [[Category: Schreuder, H A.]] |
| [[Category: Oxidoreductase]] | | [[Category: Oxidoreductase]] |
| ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Fri May 2 11:29:14 2008'' | | |
| | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Jun 30 19:07:27 2008'' |