The isolation of homogeneous leader peptidase from a strain of Escherichia coli which overproduces the enzyme.
Leader (signal) peptidases cleave the NH2-terminal leader sequences of newly synthesized secreted and membrane proteins during, or shortly after, they insert across the membrane. We have constructed a plasmid, pPS9, in which the structural gene for Escherichia coli leader peptidase is under transcriptional control of a lambda promoter, PR. pPS9 also codes for a temperature-sensitive lambda repressor, causing repression of expression of the plasmid leader peptidase gene at 30 degrees C and rapid synthesis of this enzyme at 42 degrees C. Under appropriate growth regimens, leader peptidase is overproduced 75-fold. Leader peptidase from this strain has been purified to homogeneity. It consists of a 37,000-dalton polypeptide which co-migrates with enzyme activity on DEAE-cellulose and chromatofocusing chromatography. Antibody to this leader peptidase is described.
D004262:DNA Restriction Enzymes; D010450:Endopeptidases / Q000235:genetics / Q000302:isolation & purification*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005796:Genes; D005779:Immunodiffusion; D008565:Membrane Proteins; D008970:Molecular Weight; D009876:Operon; D010957:Plasmids; D012697:Serine Endopeptidases; D013045:Species Specificity; D013696:Temperature; D014158:Transcription, Genetic
D008565:Membrane Proteins; D004262:DNA Restriction Enzymes; D010450:Endopeptidases; D012697:Serine Endopeptidases; C026515:type I signal peptidase