title: Insertion mutagenesis of the lac repressor and its implications for structure-function analysis.
abstract: We recently developed a simple technique for the generation of relatively large (31-codon) insertion mutations in cloned genes. To test whether the analysis of such mutations could provide insight into structure-function relationships in proteins, we examined a set of insertion mutants of the Escherichia coli lac repressor (LacI). Representatives of several LacI mutant classes were recovered, including mutants which exhibit fully active, inducer-insensitive, or weak dominant-negative phenotypes. The various properties of the recovered mutants agree with previous biophysical, biochemical, and genetic data for the protein. In particular, the results support the prior designation of mutationally tolerant spacer regions of LacI as well as proposed differences in dimerization interactions among regions of the protein core domain. These findings suggest that the analysis of 31-codon insertion mutations may provide a simple approach for characterizing structure-function relationships in proteins for which high-resolution structures are not available.
authors.affiliation: Department of Microbiology, University of Washington, Seattle 98195, USA.
authors.forename: B D
authors.initials: BD
authors.lastname: Nelson
authors.forename: C
authors.initials: C
authors.lastname: Manoil
authors.forename: B
authors.initials: B
authors.lastname: Traxler
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D056968:Lac Repressors; D016254:Mutagenesis, Insertional; D012097:Repressor Proteins / Q000235:genetics*; D013329:Structure-Activity Relationship
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D056968:Lac Repressors; C539064:LacI protein, E coli; D012097:Repressor Proteins