TRACKING THE EVOLUTION OF THE BACTERIAL CHOLINE-BINDING DOMAIN - MOLECULAR CHARACTERIZATION OF THE CLOSTRIDIUM-ACETOBUTYLICUM NCIB-8052 CSPA GENE

The major secreted protein of Clostridium acetobutylicum NCIB 8052, a choline-containing strain, is CspA (clostridial secreted protein). It appears to be a 115,000-M(r) glycoprotein that specifically recognizes the choline residues of the cell wall, Polyclonal antibodies raised a gainst CspA detected the presence of the protein in the cell envelope and in the culture medium, The soluble CspA protein has been purified, and an oligonucleotide probe, prepared from the determined N-terminal sequence, has been used to clone the cspA gene which encodes a protei n with 590 amino acids and an M(r) of 63,740. According to the predict ed amino acid sequence, CspA is synthesized with an N-terminal segment of 26 amino acids characteristic of prokaryotic signal peptides, Expr ession of the cspA gene in Escherichia coli led to the production of a major anti-Cspa-labeled protein of 80,000 Da which was purified by af finity chromatography on DEAE-cellulose. A comparison of CspA with oth er proteins in the EMBL database revealed that the C-terminal half of CspA is homologous to the choline-binding domains of the major pneumoc occal autolysin (LytA amidase), the pneumococcal antigen PspA, and oth er cell wall-lytic enzymes of pneumococcal phages. This region, which is constructed of four repeating motifs, also displays a high similari ty,vith the glucan-binding domains of several streptococcal glycosyltr ansferases and the toxins of Clostridium difficile.