Amino acid sequence and length requirements for assembly and function of the colicin A lysis protein.
The roles of the various parts of the mature colicin A lysis protein (Cal) in its assembly into the envelope and its function in causing "quasi-lysis," the release of colicin A, and the activation of phospholipase A were investigated. By using cassette mutagenesis, many missense mutations were introduced into the highly conserved portion of the lysis protein. In vitro mutagenesis was also used to introduce stop codons after amino acids 16 and 18 and a frameshift mutation at amino acid 17 of the mature Cal sequence. The processing and modification of the mutants were identical to those of the wild type, except for the truncated Cal proteins, which were neither acylated nor processed. Thus, the carboxy-terminal half of Cal must be present (or replaced by another peptide) for the proper processing and assembly of the protein. However, the specific sequence of this region is not required for the membrane-damaging function of the protein. Furthermore, the sequence specificity for even the conserved amino acids of the amino-terminal half of the protein is apparently exceedingly relaxed, since only those mutant Cal proteins in which a highly conserved amino acid has been replaced by a glutamate were impaired in their function.
Howard
S P
SP
Centre de Biochimie et de Biologie Moléculaire du Centre National de la Recherche Scientifique, Marseille, France.
Cavard
D
D
Lazdunski
C
C
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D003062:Codon; D004274:DNA, Recombinant / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005798:Genes, Bacterial; D007700:Kinetics; D008074:Lipoproteins; D008969:Molecular Sequence Data; D009154:Mutation; D015345:Oligonucleotide Probes; D013045:Species Specificity
D001426:Bacterial Proteins; C058485:CAL protein, Citrobacter freundii; D003062:Codon; D004274:DNA, Recombinant; D008074:Lipoproteins; D015345:Oligonucleotide Probes