Linked Life Data

1367624

Source:http://linkedlifedata.com/resource/pubmed/id/1367624

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
11
pubmed:dateCreated
1991-12-10
pubmed:abstractText
Enzyme activity is often dramatically reduced in polar organic solvents, even under conditions where the folded structures are stable. We have utilized random mutagenesis by polymerase chain reaction (PCR) techniques combined with screening for enhanced activity in the presence of dimethylformamide (DMF) to probe mechanisms by which improved enzymes for chemical synthesis in polar organic media might be obtained. Two amino acid substitutions which enhance subtilisin E activity in the presence of DMF, Q103R and D60N, were identified by screening on agar plates containing DMF and casein. The two substitutions are located near the substrate binding pocket or in the active site, and their effects on the catalytic efficiency kcat/KM for the hydrolysis of a peptide substrate are additive. The effects of D60N are apparent only in the presence of DMF, highlighting the importance of screening in the organic solvent. Protein engineering is an effective approach to enhancing enzyme activity in organic media: the triple mutant D60N + Q103R + N218S is 38 times more active than wild-type subtilisin E in 85% DMF. An evolutionary approach consisting of multiple steps of random mutagenesis and screening in continually higher concentrations of organic solvent should result in enzymes that are substantially more active in organic media.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
B
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Nov
pubmed:issn
0733-222X
pubmed:author
pubmed:issnType
Print
pubmed:volume
9
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
1073-7
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
1991
pubmed:articleTitle
Enzyme engineering for nonaqueous solvents: random mutagenesis to enhance activity of subtilisin E in polar organic media.
pubmed:affiliation
Division of Chemistry and Chemical Engineering 210-41, California Institute of Technology, Pasadena 91125.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, Non-P.H.S.