1mdr
From Proteopedia
(New page: 200px<br /><applet load="1mdr" size="450" color="white" frame="true" align="right" spinBox="true" caption="1mdr, resolution 2.1Å" /> '''THE ROLE OF LYSINE 16...) |
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| - | [[Image:1mdr.gif|left|200px]]<br /><applet load="1mdr" size=" | + | [[Image:1mdr.gif|left|200px]]<br /><applet load="1mdr" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1mdr, resolution 2.1Å" /> | caption="1mdr, resolution 2.1Å" /> | ||
'''THE ROLE OF LYSINE 166 IN THE MECHANISM OF MANDELATE RACEMASE FROM PSEUDOMONAS PUTIDA: MECHANISTIC AND CRYSTALLOGRAPHIC EVIDENCE FOR STEREOSPECIFIC ALKYLATION BY (R)-ALPHA-PHENYLGLYCIDATE'''<br /> | '''THE ROLE OF LYSINE 166 IN THE MECHANISM OF MANDELATE RACEMASE FROM PSEUDOMONAS PUTIDA: MECHANISTIC AND CRYSTALLOGRAPHIC EVIDENCE FOR STEREOSPECIFIC ALKYLATION BY (R)-ALPHA-PHENYLGLYCIDATE'''<br /> | ||
==Overview== | ==Overview== | ||
| - | The mechanism of irreversible inactivation of mandelate racemase (MR) from | + | The mechanism of irreversible inactivation of mandelate racemase (MR) from Pseudomonas putida by alpha-phenylglycidate (alpha PGA) has been investigated stereochemically and crystallographically. The (R) and (S) enantiomers of alpha PGA were synthesized in high enantiomeric excess (81% ee and 83% ee, respectively) using Sharpless epoxidation chemistry. (R)-alpha PGA was determined to be a stereospecific and stoichiometric irreversible inactivator of MR. (S)-alpha PGA does not inactivate MR and appears to bind noncovalently to the active site of MR with less affinity than that of (R)-alpha PGA. The X-ray crystal structure (2.0-A resolution) of MR inactivated by (R)-alpha PGA revealed the presence of a covalent adduct formed by nucleophilic attack of the epsilon-amino group of Lys 166 on the distal carbon on the epoxide ring of (R)-alpha PGA. The proximity of the alpha-proton of (S)-mandelate to Lys 166 [configurationally equivalent to (R)-alpha PGA] was corroborated by the crystal structure (2.1-A resolution) of MR complexed with the substrate analog/competitive inhibitor, (S)-atrolactate [(S)-alpha-methylmandelate]. These results support the proposal that Lys 166 is the polyvalent acid/base responsible for proton transfers on the (S) face of mandelate. In addition, the high-resolution structures also provide insight into the probable interactions of mandelate with the essential Mg2+ and functional groups in the active site. |
==About this Structure== | ==About this Structure== | ||
| - | 1MDR is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Pseudomonas_putida Pseudomonas putida] with MG and APG as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Mandelate_racemase Mandelate racemase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=5.1.2.2 5.1.2.2] Full crystallographic information is available from [http:// | + | 1MDR is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Pseudomonas_putida Pseudomonas putida] with <scene name='pdbligand=MG:'>MG</scene> and <scene name='pdbligand=APG:'>APG</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Mandelate_racemase Mandelate racemase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=5.1.2.2 5.1.2.2] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1MDR OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Single protein]] | [[Category: Single protein]] | ||
[[Category: Fujita, S.]] | [[Category: Fujita, S.]] | ||
| - | [[Category: Gerlt, J | + | [[Category: Gerlt, J A.]] |
| - | [[Category: Kenyon, G | + | [[Category: Kenyon, G L.]] |
| - | [[Category: Koo, C | + | [[Category: Koo, C W.]] |
| - | [[Category: Kozarich, J | + | [[Category: Kozarich, J W.]] |
| - | [[Category: Landro, J | + | [[Category: Landro, J A.]] |
| - | [[Category: Neidhart, D | + | [[Category: Neidhart, D J.]] |
| - | [[Category: Petsko, G | + | [[Category: Petsko, G A.]] |
| - | [[Category: Shah, V | + | [[Category: Shah, V J.]] |
[[Category: APG]] | [[Category: APG]] | ||
[[Category: MG]] | [[Category: MG]] | ||
[[Category: racemase]] | [[Category: racemase]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 13:54:12 2008'' |
Revision as of 11:54, 21 February 2008
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THE ROLE OF LYSINE 166 IN THE MECHANISM OF MANDELATE RACEMASE FROM PSEUDOMONAS PUTIDA: MECHANISTIC AND CRYSTALLOGRAPHIC EVIDENCE FOR STEREOSPECIFIC ALKYLATION BY (R)-ALPHA-PHENYLGLYCIDATE
Overview
The mechanism of irreversible inactivation of mandelate racemase (MR) from Pseudomonas putida by alpha-phenylglycidate (alpha PGA) has been investigated stereochemically and crystallographically. The (R) and (S) enantiomers of alpha PGA were synthesized in high enantiomeric excess (81% ee and 83% ee, respectively) using Sharpless epoxidation chemistry. (R)-alpha PGA was determined to be a stereospecific and stoichiometric irreversible inactivator of MR. (S)-alpha PGA does not inactivate MR and appears to bind noncovalently to the active site of MR with less affinity than that of (R)-alpha PGA. The X-ray crystal structure (2.0-A resolution) of MR inactivated by (R)-alpha PGA revealed the presence of a covalent adduct formed by nucleophilic attack of the epsilon-amino group of Lys 166 on the distal carbon on the epoxide ring of (R)-alpha PGA. The proximity of the alpha-proton of (S)-mandelate to Lys 166 [configurationally equivalent to (R)-alpha PGA] was corroborated by the crystal structure (2.1-A resolution) of MR complexed with the substrate analog/competitive inhibitor, (S)-atrolactate [(S)-alpha-methylmandelate]. These results support the proposal that Lys 166 is the polyvalent acid/base responsible for proton transfers on the (S) face of mandelate. In addition, the high-resolution structures also provide insight into the probable interactions of mandelate with the essential Mg2+ and functional groups in the active site.
About this Structure
1MDR is a Single protein structure of sequence from Pseudomonas putida with and as ligands. Active as Mandelate racemase, with EC number 5.1.2.2 Full crystallographic information is available from OCA.
Reference
The role of lysine 166 in the mechanism of mandelate racemase from Pseudomonas putida: mechanistic and crystallographic evidence for stereospecific alkylation by (R)-alpha-phenylglycidate., Landro JA, Gerlt JA, Kozarich JW, Koo CW, Shah VJ, Kenyon GL, Neidhart DJ, Fujita S, Petsko GA, Biochemistry. 1994 Jan 25;33(3):635-43. PMID:8292591
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