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  Structure-based prediction of modifications in glutarylamidase to allow single-step enzymatic production of 7-aminocephalosporanic acid from cephalosporin C

Fritz-Wolf, K., Koller, K.-P., Lange, G., Liesum, A., Sauber, K., Schreuder, H., et al. (2002). Structure-based prediction of modifications in glutarylamidase to allow single-step enzymatic production of 7-aminocephalosporanic acid from cephalosporin C. Protein Science, 11(1), 92-103. doi:10.1110/ps.27502.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0029-3132-6 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0029-3133-4
Genre: Journal Article
Alternative Title : Structure-based prediction of modifications in glutarylamidase to allow single-step enzymatic production of 7-aminocephalosporanic acid from cephalosporin C

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Fritz-Wolf, Karin1, Author              
Koller, Klaus-Peter, Author
Lange, Gudrun, Author
Liesum, Alexander, Author
Sauber, Klaus, Author
Schreuder, Herman, Author
Aretz, Werner, Author
Kabsch, Wolfgang1, Author              
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1Emeritus Group Biophysics, Max Planck Institute for Medical Research, Max Planck Society, ou_1497712              

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Free keywords: Cephalosporin acylase; glutaryl acylase; cephalosporin C; catalytic triad; Ntn-hydrolase; X-ray structure
 Abstract: Glutarylamidase is an important enzyme employed in the commercial production of 7-aminocephalosporanic acid, a starting compound in the synthesis of cephalosporin antibiotics. 7-aminocephalosporanic acid is obtained from cephalosporin C, a natural antibiotic, either chemically or by a two-step enzymatic process utilizing the enzymes D-amino acid oxidase and glutarylamidase. We have investigated possibilities for redesigning glutarylamidase for the production of 7-aminocephalosporanic acid from cephalosporin C in a single enzymatic step. These studies are based on the structures of glutarylamidase, which we have solved with bound phosphate and ethylene glycol to 2.5 Å resolution and with bound glycerol to 2.4 Å. The phosphate binds near the catalytic serine in a way that mimics the hemiacetal that develops during catalysis, while the glycerol occupies the side-chain binding pocket. Our structures show that the enzyme is not only structurally similar to penicillin G acylase but also employs essentially the same mechanism in which the α-amino group of the catalytic serine acts as a base. A subtle difference is the presence of two catalytic dyads, His B23/Glu B455 and His B23/Ser B1, that are not seen in penicillin G acylase. In contrast to classical serine proteases, the central histidine of these dyads interacts indirectly with the Oγ through a hydrogen bond relay network involving the α-amino group of the serine and a bound water molecule. A plausible model of the enzyme–substrate complex is proposed that leads to the prediction of mutants of glutarylamidase that should enable the enzyme to deacylate cephalosporin C into 7-aminocephalosporanic acid.

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Language(s): eng - English
 Dates: 20012001-10-102009-04-132002-01-01
 Publication Status: Published in print
 Pages: 12
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 Table of Contents: -
 Rev. Type: Peer
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Title: Protein Science
Source Genre: Journal
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Publ. Info: New York, N.Y. : Cambridge University Press
Pages: - Volume / Issue: 11 (1) Sequence Number: - Start / End Page: 92 - 103 Identifier: ISSN: 0961-8368
CoNE: https://pure.mpg.de/cone/journals/resource/954925342760