2puh
From Proteopedia
| Line 1: | Line 1: | ||
| - | [[Image:2puh.jpg|left|200px]] | + | [[Image:2puh.jpg|left|200px]] |
| - | + | ||
| - | '''Crystal Structure of the S112A mutant of a C-C hydrolase, BphD from Burkholderia xenovorans LB400, in complex with its substrate HOPDA''' | + | {{Structure |
| + | |PDB= 2puh |SIZE=350|CAPTION= <scene name='initialview01'>2puh</scene>, resolution 1.820Å | ||
| + | |SITE= | ||
| + | |LIGAND= <scene name='pdbligand=NA:SODIUM+ION'>NA</scene>, <scene name='pdbligand=MLI:MALONATE+ION'>MLI</scene> and <scene name='pdbligand=HPK:(3E)-2,6-DIOXO-6-PHENYLHEX-3-ENOATE'>HPK</scene> | ||
| + | |ACTIVITY= | ||
| + | |GENE= bphD ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=36873 Burkholderia xenovorans]) | ||
| + | }} | ||
| + | |||
| + | '''Crystal Structure of the S112A mutant of a C-C hydrolase, BphD from Burkholderia xenovorans LB400, in complex with its substrate HOPDA''' | ||
| + | |||
==Overview== | ==Overview== | ||
| Line 7: | Line 16: | ||
==About this Structure== | ==About this Structure== | ||
| - | 2PUH is a [ | + | 2PUH is a [[Single protein]] structure of sequence from [http://en.wikipedia.org/wiki/Burkholderia_xenovorans Burkholderia xenovorans]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2PUH OCA]. |
==Reference== | ==Reference== | ||
| - | The tautomeric half-reaction of BphD, a C-C bond hydrolase. Kinetic and structural evidence supporting a key role for histidine 265 of the catalytic triad., Horsman GP, Bhowmik S, Seah SY, Kumar P, Bolin JT, Eltis LD, J Biol Chem. 2007 Jul 6;282(27):19894-904. Epub 2007 Apr 18. PMID:[http:// | + | The tautomeric half-reaction of BphD, a C-C bond hydrolase. Kinetic and structural evidence supporting a key role for histidine 265 of the catalytic triad., Horsman GP, Bhowmik S, Seah SY, Kumar P, Bolin JT, Eltis LD, J Biol Chem. 2007 Jul 6;282(27):19894-904. Epub 2007 Apr 18. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/17442675 17442675] |
[[Category: Burkholderia xenovorans]] | [[Category: Burkholderia xenovorans]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| Line 20: | Line 29: | ||
[[Category: c-c bond hydrolase]] | [[Category: c-c bond hydrolase]] | ||
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Mar 20 18:17:31 2008'' |
Revision as of 16:17, 20 March 2008
| |||||||
| , resolution 1.820Å | |||||||
|---|---|---|---|---|---|---|---|
| Ligands: | , and | ||||||
| Gene: | bphD (Burkholderia xenovorans) | ||||||
| Coordinates: | save as pdb, mmCIF, xml | ||||||
Crystal Structure of the S112A mutant of a C-C hydrolase, BphD from Burkholderia xenovorans LB400, in complex with its substrate HOPDA
Overview
BphD of Burkholderia xenovorans LB400 catalyzes an unusual C-C bond hydrolysis of 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid (HOPDA) to afford benzoic acid and 2-hydroxy-2,4-pentadienoic acid (HPD). An enol-keto tautomerization has been proposed to precede hydrolysis via a gem-diol intermediate. The role of the canonical catalytic triad (Ser-112, His-265, Asp-237) in mediating these two half-reactions remains unclear. We previously reported that the BphD-catalyzed hydrolysis of HOPDA (lambda(max) is 434 nm for the free enolate) proceeds via an unidentified intermediate with a red-shifted absorption spectrum (lambda(max) is 492 nm) (Horsman, G. P., Ke, J., Dai, S., Seah, S. Y. K., Bolin, J. T., and Eltis, L. D. (2006) Biochemistry 45, 11071-11086). Here we demonstrate that the S112A variant generates and traps a similar intermediate (lambda(max) is 506 nm) with a similar rate, 1/tau approximately 500 s(-1). The crystal structure of the S112A:HOPDA complex at 1.8-A resolution identified this intermediate as the keto tautomer, (E)-2,6-dioxo-6-phenyl-hex-3-enoate. This keto tautomer did not accumulate in either the H265A or the S112A/H265A double variants, indicating that His-265 catalyzes tautomerization. Consistent with this role, the wild type and S112A enzymes catalyzed tautomerization of the product HPD, whereas H265A variants did not. This study thus identifies a keto intermediate, and demonstrates that the catalytic triad histidine catalyzes the tautomerization half-reaction, expanding the role of this residue from its purely hydrolytic function in other serine hydrolases. Finally, the S112A:HOPDA crystal structure is more consistent with hydrolysis occurring via an acyl-enzyme intermediate than a gem-diol intermediate as solvent molecules have poor access to C6, and the closest ordered water is 7 A away.
About this Structure
2PUH is a Single protein structure of sequence from Burkholderia xenovorans. Full crystallographic information is available from OCA.
Reference
The tautomeric half-reaction of BphD, a C-C bond hydrolase. Kinetic and structural evidence supporting a key role for histidine 265 of the catalytic triad., Horsman GP, Bhowmik S, Seah SY, Kumar P, Bolin JT, Eltis LD, J Biol Chem. 2007 Jul 6;282(27):19894-904. Epub 2007 Apr 18. PMID:17442675
Page seeded by OCA on Thu Mar 20 18:17:31 2008
