A gene (sleC) encoding a spore-cortex-lytic enzyme from Clostridium perfringens S40 spores; cloning, sequence analysis and molecular characterization.
Antiserum was raised against a 31 kDa spore-cortex-lytic enzyme, which is released during germination of Clostridium perfringens S40 spores. Western blotting of dormant spore and vegetative cell fractions separated by SDS-PAGE indicated that the 31 kDa enzyme is spore-specific and that the enzyme in the dormant spore exists as a 36 kDa protein which has no cortex-lytic activity. A gene encoding the 31 kDa enzyme, sleC, was cloned into Escherichia coli using a synthetic oligonucleotide as a hybridization probe and the nucleotide sequence of the entire gene was determined. The N-terminal amino acid sequence of the 36 kDa protein was found in this reading frame, confirming that the 36 kDa protein is a pro-form of the 31 kDa enzyme. The deduced amino acid sequence indicated that the 31 kDa enzyme is produced as a precursor, comprising three portions; an N-terminal prepro-sequence (114 amino acid residues), a pro-sequence (35 amino acid residues) and a mature enzyme (289 amino acid residues). It is suggested that the 36 kDa pro-enzyme is non-covalently attached to the exterior of the cortex layer, and that the proform is processed to release the active enzyme during germination.
Miyata
S
S
Department of Applied Biological Sciences, School of Agricultural Sciences, Nagoya University, Japan.
Moriyama
R
R
Miyahara
N
N
Makino
S
S
D000595:Amino Acid Sequence; D001433:Bacteriolysis; D001483:Base Sequence; D015153:Blotting, Western; D002451:Cell Compartmentation; D003001:Cloning, Molecular; D003016:Clostridium perfringens / Q000201:enzymology / Q000235:genetics*; D003429:Cross Reactions; D004792:Enzyme Precursors / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D015698:Genomic Library; D008969:Molecular Sequence Data; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000235:genetics* / Q000276:immunology; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D013171:Spores, Bacterial / Q000201:enzymology / Q000235:genetics
D004792:Enzyme Precursors; D009238:N-Acetylmuramoyl-L-alanine Amidase