Cloning and expression of gene fragment IgA-binding protein of group B streptococci.
Different fragments of the bac gene coding for the IgA-binding protein were cloned, sequenced and expressed in E. 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 bp bac 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.
Ustinovitch
I
I
Institute of Experimental Medicine, Russian Academy of Medical Sciences, St. Petersburg, Russia.
Vlasov
G
G
Totolyan
A
A
Suvorov
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D007070:Immunoglobulin A / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D011817:Rabbits; D013292:Streptococcus agalactiae / Q000235:genetics* / Q000276:immunology / Q000378:metabolism
D017931:DNA Primers; D004268:DNA-Binding Proteins; D007070:Immunoglobulin A; C082525:beta protein, Streptococcus