title: Genetic analysis of the LexA repressor: isolation and characterization of LexA(Def) mutant proteins.
abstract: We report the isolation of LexA mutant proteins with impaired repressor function. These mutant proteins were obtained by transforming a LexA-deficient recA-lacZ indicator strain with a randomly mutagenized plasmid harbouring the lexA gene and subsequent selection on MacConkey-lactose indicator plates. A total of 24 different lexA(Def) missense mutations were identified. All except three mutant proteins are produced in near-normal amounts suggesting that they are fairly resistant to intracellular proteases. All lexA(Def) missense mutations are situated within the first 67 amino acids of the amino-terminal DNA binding domain. The properties of an intragenic deletion mutant suggest that the part of the amino-terminal domain important for DNA recognition or domain folding should extent at least to amino acids 69 or 70. A recent 2D-NMR study (Lamerichs et al. 1989) has identified three alpha helices in the DNA binding domain of LexA. The relative orientation of two of them (helices 2 and 3) is reminiscent of, but not identical to, the canonical helix-turn-helix motif suggesting nevertheless that helix 3 might be involved in DNA recognition. The distribution of the lexA(Def) missense mutations along the first 67 amino-terminal amino acids indeed shows some clustering within helix 3, since 8 out of the 24 different missense mutations are found in this helix. However one mutation in front of helix 1 and five mutations between amino acids 61 and 67 suggest that elements other than helices 2 and 3 may be important for DNA binding.
authors: Institut de Biologie Moléculaire et Cellulaire, CNRS-LP 6201, INSERM, Strasbourg, France.
authors: P
authors: P
authors: Oertel-Buchheit
authors: R M
authors: RM
authors: Lamerichs
authors: M
authors: M
authors: Schnarr
authors: M
authors: M
authors: Granger-Schnarr
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D003062:Codon; D015151:Immunoblotting; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000235:genetics*; D012697:Serine Endopeptidases; D014170:Transformation, Genetic; D000099160:LexA Repressor Protein
chemical_list: D001426:Bacterial Proteins; D003062:Codon; D012097:Repressor Proteins; D012697:Serine Endopeptidases; D000099160:LexA Repressor Protein