Cloning and expression of gene fragment IgA-binding protein of group B streptococci

Abstract

Different fragments of thebac gene coding for the IgA-binding protein were cloned, sequenced and expressed inE. coli. Cloning was accomplished after amplification of different parts of the gene by PCR. The 1.5-kb fragment of the gene was cloned using plasmid pBluescript. This fragment coded for the 45-kDa protein with the stable expression of IgA binding. In order to verify the exact location of the IgA-binding domain two smaller plasmids were constructed. Both plasmids were prepared using pQE30 (31, 32) expression vectors from Qiagen. The plasmids carried 245 and 123 bpbac gene fragments encoding 14-and 7-kDa proteins. These proteins together with the 20-amino-acid oligopeptide ITNEDKDSMLKKIEDINRQA were tested for IgA binding. Only the 14-kDa protein was able to bind IgA. This protein was used for rabbit immunization and found to be immunogenic. The data obtained lead to the conclusion that there is a lower limit in the size of recombinant IgA-binding proteins that can be utilized for anti-GBS vaccination.