1zk3
From Proteopedia
(New page: 200px<br /><applet load="1zk3" size="450" color="white" frame="true" align="right" spinBox="true" caption="1zk3, resolution 2.2Å" /> '''Triclinic crystal str...) |
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| - | [[Image:1zk3.gif|left|200px]]<br /><applet load="1zk3" size=" | + | [[Image:1zk3.gif|left|200px]]<br /><applet load="1zk3" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1zk3, resolution 2.2Å" /> | caption="1zk3, resolution 2.2Å" /> | ||
'''Triclinic crystal structure of the apo-form of R-specific alcohol dehydrogenase (mutant G37D) from Lactobacillus brevis'''<br /> | '''Triclinic crystal structure of the apo-form of R-specific alcohol dehydrogenase (mutant G37D) from Lactobacillus brevis'''<br /> | ||
==Overview== | ==Overview== | ||
| - | The R-specific alcohol dehydrogenase (RADH) from Lactobacillus brevis is | + | The R-specific alcohol dehydrogenase (RADH) from Lactobacillus brevis is an NADP-dependent, homotetrameric member of the extended enzyme family of short-chain dehydrogenases/reductases (SDR) with a high biotechnological application potential. Its preferred in vitro substrates are prochiral ketones like acetophenone with almost invariably a small methyl group as one substituent and a bulky (often aromatic) moiety as the other. On the basis of an atomic-resolution structure of wild-type RADH in complex with NADP and acetophenone, we designed the mutant RADH-G37D, which should possess an improved cosubstrate specificity profile for biotechnological purposes, namely, a preference for NAD rather than NADP. Comparative kinetic measurements with wild-type and mutant RADH showed that this aim was achieved. To characterize the successful mutant structurally, we determined several, partly atomic-resolution, crystal structures of RADH-G37D both as an apo-enzyme and as ternary complex with NAD or NADH and phenylethanol. The increased affinity of RADH-G37D for NAD(H) depends on an interaction between the adenosine ribose moiety of NAD and the inserted aspartate side-chain. A structural comparison between RADH-G37D as apo-enzyme and as a part of a ternary complex revealed significant rearrangements of Ser141, Glu144, Tyr189 and Met205 in the vicinity of the active site. This plasticity contributes to generate a small hydrophobic pocket for the methyl group typical for RADH substrates, and a hydrophobic coat for the second, more variable and often aromatic, substituent. Around Ser141 we even found alternative conformations in the backbone. A structural adaptability in this region, which we describe here for the first time for an SDR enzyme, is probably functionally important, because it concerns Ser142, a member of the highly conserved catalytic tetrad typical for SDR enzymes. Moreover, it affects an extended proton relay system that has been identified recently as a critical element for the catalytic mechanism in SDR enzymes. |
==About this Structure== | ==About this Structure== | ||
| - | 1ZK3 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Lactobacillus_brevis Lactobacillus brevis] with MG as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Alcohol_dehydrogenase_(NADP(+)) Alcohol dehydrogenase (NADP(+))], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.1.1.2 1.1.1.2] Full crystallographic information is available from [http:// | + | 1ZK3 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Lactobacillus_brevis Lactobacillus brevis] with <scene name='pdbligand=MG:'>MG</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Alcohol_dehydrogenase_(NADP(+)) Alcohol dehydrogenase (NADP(+))], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.1.1.2 1.1.1.2] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1ZK3 OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Niefind, K.]] | [[Category: Niefind, K.]] | ||
[[Category: Riebel, B.]] | [[Category: Riebel, B.]] | ||
| - | [[Category: Schlieben, N | + | [[Category: Schlieben, N H.]] |
[[Category: Schomburg, D.]] | [[Category: Schomburg, D.]] | ||
[[Category: MG]] | [[Category: MG]] | ||
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[[Category: short chain reductases/dehydrogenases]] | [[Category: short chain reductases/dehydrogenases]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:16:21 2008'' |
Revision as of 14:16, 21 February 2008
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Triclinic crystal structure of the apo-form of R-specific alcohol dehydrogenase (mutant G37D) from Lactobacillus brevis
Overview
The R-specific alcohol dehydrogenase (RADH) from Lactobacillus brevis is an NADP-dependent, homotetrameric member of the extended enzyme family of short-chain dehydrogenases/reductases (SDR) with a high biotechnological application potential. Its preferred in vitro substrates are prochiral ketones like acetophenone with almost invariably a small methyl group as one substituent and a bulky (often aromatic) moiety as the other. On the basis of an atomic-resolution structure of wild-type RADH in complex with NADP and acetophenone, we designed the mutant RADH-G37D, which should possess an improved cosubstrate specificity profile for biotechnological purposes, namely, a preference for NAD rather than NADP. Comparative kinetic measurements with wild-type and mutant RADH showed that this aim was achieved. To characterize the successful mutant structurally, we determined several, partly atomic-resolution, crystal structures of RADH-G37D both as an apo-enzyme and as ternary complex with NAD or NADH and phenylethanol. The increased affinity of RADH-G37D for NAD(H) depends on an interaction between the adenosine ribose moiety of NAD and the inserted aspartate side-chain. A structural comparison between RADH-G37D as apo-enzyme and as a part of a ternary complex revealed significant rearrangements of Ser141, Glu144, Tyr189 and Met205 in the vicinity of the active site. This plasticity contributes to generate a small hydrophobic pocket for the methyl group typical for RADH substrates, and a hydrophobic coat for the second, more variable and often aromatic, substituent. Around Ser141 we even found alternative conformations in the backbone. A structural adaptability in this region, which we describe here for the first time for an SDR enzyme, is probably functionally important, because it concerns Ser142, a member of the highly conserved catalytic tetrad typical for SDR enzymes. Moreover, it affects an extended proton relay system that has been identified recently as a critical element for the catalytic mechanism in SDR enzymes.
About this Structure
1ZK3 is a Single protein structure of sequence from Lactobacillus brevis with as ligand. Active as Alcohol dehydrogenase (NADP(+)), with EC number 1.1.1.2 Full crystallographic information is available from OCA.
Reference
Atomic resolution structures of R-specific alcohol dehydrogenase from Lactobacillus brevis provide the structural bases of its substrate and cosubstrate specificity., Schlieben NH, Niefind K, Muller J, Riebel B, Hummel W, Schomburg D, J Mol Biol. 2005 Jun 17;349(4):801-13. PMID:15896805
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