Linked Life Data

12082164

Source:http://linkedlifedata.com/resource/pubmed/id/12082164

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
6
pubmed:dateCreated
2002-6-25
pubmed:abstractText
Circular permutation of proteins is a powerful technique to explore the importance of the polypeptide secondary structure order for attaining the final three-dimensional structure. Here, we designed a circular permutation of the TEM beta-lactamase in order to produce a new domain-forming amino acid arrangement in the polypeptide sequence. Closing the normal N- and C-termini with the connecting peptide GGS and creating new N- and C-termini at position 216, produces a severely impaired permuted protein. Introduction of a connector with random components allows the isolation of enzymes with better activities and indicates a selection for a potential helix-stop signal at the new super-secondary motif. We applied several directed-evolution cycles, starting from permuted enzymes with each of the two different connecting peptides, and selecting for antibiotic resistance and isolated several mutants with resistance levels close to those of the wild-type enzyme. We also analyze some of the data collected on the outcomes and paths of these evolutionary experiments. A purified sixth cycle variant with connector peptide GGS showed catalytic efficiency values approximately 8% of the natural enzyme.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
0269-2139
pubmed:author
pubmed:issnType
Print
pubmed:volume
15
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
463-70
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed:year
2002
pubmed:articleTitle
Improving a circularly permuted TEM-1 beta-lactamase by directed evolution.
pubmed:affiliation
Instituto de Biotecnología, UNAM, Apdo. Postal 510-3 Cuernavaca, Morelos 62250, México. joel@ibt.unam.mx
pubmed:publicationType
Journal Article, In Vitro, Research Support, Non-U.S. Gov't