Linked Life Data

10712737

Source:http://linkedlifedata.com/resource/pubmed/id/10712737

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2000-6-20
pubmed:abstractText
Bifunctional enzymes find a wide application as a monitoring facility and a potential biocatalyst in molecular biology and biotechnology. Recombination of natural enzymes to a bifunctional fusion offers valuable tools, but the functional and structural instability of artificial fusion enzymes remains to be solved. Based on structural traits of microbial D-hydantoinase, we attempted to construct a bifunctional N-carbamylase/D-hydantoinase fusion enzyme that would be useful for the synthesis of nonnatural D-amino acids in a concerted fashion. The bifunctional ability of D-hydantoinase, as a fusion partner, was noticeable, but the resulting fusion enzyme was subjected to serious proteolysis in vivo, as generally encountered in the expression of large the multidomain polypeptide in E. coli. In an effort to improve the structural instability imposed by artificial linear fusion, directed evolution of the fusion enzyme was performed using DNA shuffling with a consensus primer to maintain a crucial domain for the enzyme activity. The evolved fusion enzyme, F11, was selected after repeated rounds, and this enzyme was found to show sixfold increased performance in the production of D-amino acid compared with the parent fusion enzyme, which was mainly due to the enhanced structural stability of the evolved fusion enzyme. This result is an example showing that directed evolution of the linearly fused polypeptide may broaden the opportunity to generate a fusion enzyme with greater potential.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Apr
pubmed:issn
0006-3592
pubmed:author
pubmed:copyrightInfo
Copyright 2000 John Wiley & Sons, Inc.
pubmed:issnType
Print
pubmed:day
20
pubmed:volume
68
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
211-7
pubmed:dateRevised
2009-11-19
pubmed:meshHeading
pubmed:year
2000
pubmed:articleTitle
Directed evolution of a novel N-carbamylase/D-hydantoinase fusion enzyme for functional expression with enhanced stability.
pubmed:affiliation
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1, Kusung-dong, Yusung-gu, Taejon 305-701, Korea.
pubmed:publicationType
Journal Article