1ykj
From Proteopedia
(New page: 200px<br /><applet load="1ykj" size="450" color="white" frame="true" align="right" spinBox="true" caption="1ykj, resolution 2.00Å" /> '''A45G p-hydroxybenzoa...) |
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| - | [[Image:1ykj.gif|left|200px]]<br /><applet load="1ykj" size=" | + | [[Image:1ykj.gif|left|200px]]<br /><applet load="1ykj" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1ykj, resolution 2.00Å" /> | caption="1ykj, resolution 2.00Å" /> | ||
'''A45G p-hydroxybenzoate hydroxylase with p-hydroxybenzoate bound'''<br /> | '''A45G p-hydroxybenzoate hydroxylase with p-hydroxybenzoate bound'''<br /> | ||
==Overview== | ==Overview== | ||
| - | p-Hydroxybenzoate hydroxylase (PHBH) is a homodimeric flavoprotein | + | p-Hydroxybenzoate hydroxylase (PHBH) is a homodimeric flavoprotein monooxygenase that catalyzes the hydroxylation of p-hydroxybenzoate to form 3,4-dihydroxybenzoate. Controlled catalysis is achieved by movement of the flavin and protein between three conformations, in, out, and open [Entsch, B., et al. (2005) Arch. Biochem. Biophys. 433, 297-311]. The open conformation is important for substrate binding and product release, the in conformation for reaction with oxygen and hydroxylation, and the out conformation for the reduction of FAD by NADPH. The open conformation is similar to the structure of Arg220Gln-PHBH in which the backbone peptide loop of residues 43-46, located on the si side of the flavin, is rotated. In this paper, we examine the structure and properties of the Ala45Gly-PHBH mutant enzyme. The crystal structure of the Ala45Gly enzyme is an asymmetric dimer, with one monomer similar (but not identical) to wild-type PHBH, while the other monomer has His72 flipped into solvent and replaced with Glu73 as one of several changes in the structure. The two structures correlate with evidence from kinetic studies for two forms of Ala45Gly-PHBH. One form of the enzyme dominates turnover and hydroxylates, while the other contributes little to turnover and fails to hydroxylate. Ala45Gly-PHBH favors the in conformation over alternative conformations. The effect of this mutation on the structure and function of PHBH illustrates the importance of the si side loop in the conformational state of PHBH and, consequently, the function of the enzyme. This work demonstrates some general principles of how enzymes use conformational movements to allow both access and egress of substrates and product, while restricting access to the solvent at a critical stage in catalysis. |
==About this Structure== | ==About this Structure== | ||
| - | 1YKJ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Pseudomonas_aeruginosa Pseudomonas aeruginosa] with SO4, FAD, PHB and PSL as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/4-hydroxybenzoate_3-monooxygenase 4-hydroxybenzoate 3-monooxygenase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.14.13.2 1.14.13.2] Full crystallographic information is available from [http:// | + | 1YKJ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Pseudomonas_aeruginosa Pseudomonas aeruginosa] with <scene name='pdbligand=SO4:'>SO4</scene>, <scene name='pdbligand=FAD:'>FAD</scene>, <scene name='pdbligand=PHB:'>PHB</scene> and <scene name='pdbligand=PSL:'>PSL</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/4-hydroxybenzoate_3-monooxygenase 4-hydroxybenzoate 3-monooxygenase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.14.13.2 1.14.13.2] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1YKJ OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Pseudomonas aeruginosa]] | [[Category: Pseudomonas aeruginosa]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Ballou, D | + | [[Category: Ballou, D P.]] |
| - | [[Category: Cole, L | + | [[Category: Cole, L J.]] |
[[Category: Entsch, B.]] | [[Category: Entsch, B.]] | ||
| - | [[Category: Gatti, D | + | [[Category: Gatti, D L.]] |
[[Category: FAD]] | [[Category: FAD]] | ||
[[Category: PHB]] | [[Category: PHB]] | ||
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[[Category: phbh]] | [[Category: phbh]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:06:16 2008'' |
Revision as of 14:06, 21 February 2008
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A45G p-hydroxybenzoate hydroxylase with p-hydroxybenzoate bound
Overview
p-Hydroxybenzoate hydroxylase (PHBH) is a homodimeric flavoprotein monooxygenase that catalyzes the hydroxylation of p-hydroxybenzoate to form 3,4-dihydroxybenzoate. Controlled catalysis is achieved by movement of the flavin and protein between three conformations, in, out, and open [Entsch, B., et al. (2005) Arch. Biochem. Biophys. 433, 297-311]. The open conformation is important for substrate binding and product release, the in conformation for reaction with oxygen and hydroxylation, and the out conformation for the reduction of FAD by NADPH. The open conformation is similar to the structure of Arg220Gln-PHBH in which the backbone peptide loop of residues 43-46, located on the si side of the flavin, is rotated. In this paper, we examine the structure and properties of the Ala45Gly-PHBH mutant enzyme. The crystal structure of the Ala45Gly enzyme is an asymmetric dimer, with one monomer similar (but not identical) to wild-type PHBH, while the other monomer has His72 flipped into solvent and replaced with Glu73 as one of several changes in the structure. The two structures correlate with evidence from kinetic studies for two forms of Ala45Gly-PHBH. One form of the enzyme dominates turnover and hydroxylates, while the other contributes little to turnover and fails to hydroxylate. Ala45Gly-PHBH favors the in conformation over alternative conformations. The effect of this mutation on the structure and function of PHBH illustrates the importance of the si side loop in the conformational state of PHBH and, consequently, the function of the enzyme. This work demonstrates some general principles of how enzymes use conformational movements to allow both access and egress of substrates and product, while restricting access to the solvent at a critical stage in catalysis.
About this Structure
1YKJ is a Single protein structure of sequence from Pseudomonas aeruginosa with , , and as ligands. Active as 4-hydroxybenzoate 3-monooxygenase, with EC number 1.14.13.2 Full crystallographic information is available from OCA.
Reference
Removal of a methyl group causes global changes in p-hydroxybenzoate hydroxylase., Cole LJ, Gatti DL, Entsch B, Ballou DP, Biochemistry. 2005 Jun 7;44(22):8047-58. PMID:15924424
Page seeded by OCA on Thu Feb 21 16:06:16 2008
Categories: 4-hydroxybenzoate 3-monooxygenase | Pseudomonas aeruginosa | Single protein | Ballou, D P. | Cole, L J. | Entsch, B. | Gatti, D L. | FAD | PHB | PSL | SO4 | Catalysis | Conformations | Phbh
