Source:http://linkedlifedata.com/resource/pubmed/id/15333773
Switch to
| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
7
|
| pubmed:dateCreated |
2004-11-4
|
| pubmed:abstractText |
A semi-random mutagenesis approach was followed to increase the performance of penicillin acylase PAS2 in the kinetically controlled synthesis of ampicillin from 6-aminopenicillanic acid (6-APA) and activated D-phenylglycine derivatives. We directed changes in amino acid residues to positions close to the active site that are expected to affect the catalytic performance of penicillin acylase: alpha R160, alpha F161 and beta F24. From the resulting triple mutant gene bank, six improved PAS2 mutants were recovered by screening only 700 active mutants with an HPLC-based screening method. A detailed kinetic analysis of the three most promising mutants, T23, TM33 and TM38, is presented. These mutants allowed the accumulation of ampicillin at 4-5 times higher concentrations than the wild-type enzyme, using D-phenylglycine methyl ester as the acyl donor. At the same time, the loss of activated acyl donor due to the competitive hydrolytic side reactions could be reduced to <20% with the mutant enzymes compared >80% wild-type PAS2. Although catalytic activity dropped by a factor of 5-10, the enhanced synthetic performance of the recovered penicillin acylase variants makes them interesting biocatalysts for the production of beta-lactam antibiotics.
|
| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Glycine,
http://linkedlifedata.com/resource/pubmed/chemical/Penicillanic Acid,
http://linkedlifedata.com/resource/pubmed/chemical/Penicillin Amidase,
http://linkedlifedata.com/resource/pubmed/chemical/aminopenicillanic acid,
http://linkedlifedata.com/resource/pubmed/chemical/beta-Lactams,
http://linkedlifedata.com/resource/pubmed/chemical/methyl phenylglycine
|
| pubmed:status |
MEDLINE
|
| pubmed:month |
Jul
|
| pubmed:issn |
1741-0126
|
| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:volume |
17
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
571-9
|
| pubmed:dateRevised |
2006-5-1
|
| pubmed:meshHeading |
pubmed-meshheading:15333773-Catalysis,
pubmed-meshheading:15333773-Chromatography, High Pressure Liquid,
pubmed-meshheading:15333773-Dose-Response Relationship, Drug,
pubmed-meshheading:15333773-Escherichia coli,
pubmed-meshheading:15333773-Genetic Techniques,
pubmed-meshheading:15333773-Glycine,
pubmed-meshheading:15333773-Hydrolysis,
pubmed-meshheading:15333773-Kinetics,
pubmed-meshheading:15333773-Models, Chemical,
pubmed-meshheading:15333773-Mutagenesis,
pubmed-meshheading:15333773-Mutation,
pubmed-meshheading:15333773-Penicillanic Acid,
pubmed-meshheading:15333773-Penicillin Amidase,
pubmed-meshheading:15333773-Plasmids,
pubmed-meshheading:15333773-Protein Conformation,
pubmed-meshheading:15333773-Protein Engineering,
pubmed-meshheading:15333773-Time Factors,
pubmed-meshheading:15333773-beta-Lactams
|
| pubmed:year |
2004
|
| pubmed:articleTitle |
Increasing the synthetic performance of penicillin acylase PAS2 by structure-inspired semi-random mutagenesis.
|
| pubmed:affiliation |
Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
|
| pubmed:publicationType |
Journal Article
|