Cloning of a new low-molecular-weight spore-specific protein gene from Bacillus megaterium

Abstract

Three EcoRI fragments of Bacillus megaterium DNA hybridized only under nonrestrictive conditions on Southern blots to a probe containing the previously cloned gene for protein C, a small, acid-soluble spore protein (SASP) from B. megaterium. All three fragments were cloned in Escherichia coli cells in plasmid pBR325, and after being transferred to an E. coli expression vector, one of the fragments (C-3) directed the synthesis of a new small, acid-soluble spore protein (termed C-3) immunologically related to protein C. As previously observed with the protein C gene, protein C-3 gene expression in E. coli required an external promoter and suppression of termination of transcription. Protein C-3 was purified from induced E. coli cells, and its immunological properties, electrophoretic mobility, amino acid composition, and amino-terminal sequence were determined. These data indicated that protein C-3 was related, but not identical, to either protein C or the closely related protein A--two of the major small, acid-soluble spore proteins of B. megaterium. Detailed examination of acid extracts of B. megaterium spores showed that they contained a minor protein which comigrated with C-3 on acrylamide gel electrophoresis at low pH and reacted immunologically like C-3.