Crystal structure of Deinococcus radiodurans N-acylamino acid racemase at 1.3Å

• Main Authors: Chun-Li Wu, 吳峻岦
• Other Authors: W.C. Wang
• Format: Others
• Language: zh-TW
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/72271613918501336617

碩士 === 國立清華大學 === 生命科學系 === 91 === The members of the enolase superfamily sharing the common ability to catalyze abstraction of the α-proton of carboxylic acids. Although each reaction catalyzed by these enzymes is initiated by this common step, their overall reactions as well as the stereochemical consequences of the β-elimination reactions are diverse. A protein identified as “N-acylamino acid racemase”, a member of enolase superfamily, racemizes optically active N-acylamino acid in the presence of optical active amino acid; its use in combination with D- or L-aminoacylase enables the production of optically active D- or L-alpha-amino acid from DL-acyl-alpha-aminocarboxylic acid at a high level of efficiency. N-acylamino acid racemase from Amycolaptosis sp. is an inefficient enzyme（kcat/Km＝3.7×102 M-1 s-1）. Its sequence is 43% identical to that of an unidentified protein encoded by the Bacillus subtilis genome. Both proteins efficiently catalyze the o-succinylbenzoate synthase reaction in menaquinone biosynthesis（kcat/Km＝2.5×105 and 7.5×105 M-1 s-1, respectively）, suggesting that this is their “correct” metabolic function. We have solved the X-ray structure of N-acylamino acid racemase from Deinococcus radiodurans to 1.3Å resolution by multiple wavelength anomalous diffraction（MAD）. The two-domanins fold found for N-acylamino acid racemase is similar to those for other members of the enolase superfamily：a mixed α/β capping domain formed from segments at the N- and C-termini of the polypeptide and a larger（β/α）7β barrel domain. We have compared the structure with o-succinylbenzoate synthase, and speculated that Lys168，Lys170，Asp195，Glu220，Asp245，Lys269 are the key residues in the catalytic region. As a result, evolution of both the shape and volume of the active site should be subject to few structural constraints. This would provide a structural strategy for the evolution of new catalytic activities in homologues of enolase superfamily and a likely explanation for how the o-succinylbenzoate synthase from Amycolaptosis sp. also can catalyze the reaction of N-acylamino acid racemase（Palmer et al., 1999; Thomas et al., 2000; Andrew et al., 2001）.