Limited proteolysis. Early steps in the processing of large premature termination fragments of beta-galactosidase in Escherichia coli.
The mechanism by which large premature termination fragments of beta-galactosidase were degraded in Escherichia coli was studied using quantitative immunoprecipitation techniques. Two different lacZ nonsense mutants which produced apparent primary translation products of 96,000 and 109,000 daltons, respectively, were both shown to produce a second beta-galactosidase-related polypeptide of Mr = 90,000. These 90,000-dalton polypeptides appeared to be the same in both strains since they co-migrated when analyzed as a mixture on sodium dodecyl sulfate-polyacrylamide gels and were indistinguishable when analyzed by one-dimensional peptide mapping. Pulse-chase experiments established a stoichiometric precursor-product relationship between the primary mutant gene products (called the A polypeptides) and the common 90,000-dalton polypeptide (called the B polypeptide). No intermediates were detected between the A and B polypeptides. We propose that there is a common pathway for the degradation of these different large fragments of beta-galactosidase. According to this model, the first step would be a specific endoproteolytic cleavage of the primary translation product which produces the 90,000-dalton polypeptide as a common intermediate. The kinetic analysis demonstrated a first order decay of both A and B polypeptides but, surprisingly, the first order rate constant for the decay of A appeared dependent upon the induction regimen. This result suggested that degradation may possibly be autoregulated either by the intracellular level of A or by other intermediates in the degradation pathway.
D000936:Antigen-Antibody Complex; D000939:Epitopes / Q000032:analysis; D004926:Escherichia coli / Q000201:enzymology*; D005696:Galactosidases / Q000235:genetics*; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009154:Mutation; D010443:Peptide Chain Termination, Translational; D013045:Species Specificity; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics*
D000936:Antigen-Antibody Complex; D000939:Epitopes; D046911:Macromolecular Substances; D005696:Galactosidases; D001616:beta-Galactosidase