Expression and purification of the Bacillus anthracis protective antigen receptor-binding domain

Objective To express the receptor-binding domain of Bacillus anthracis protective antigen in E. coli. Methods Signal sequence of the outer membrane protein A(OmpA)of E. coli was attached to the 5′ end of the gene encoding protective antigen receptor-binding domain(the fourth domain of PA, PA-D4). The plasmid carrying the fusion gene was then transformed into E. coli and induced to express recombinant PA-D4 by IPTG. The recombinant protein was purified by chromatography and then identified by N-terminal sequencing and Western blot. Results The recombinant protein, about 10% of the total bacterial protein in volume, was secreted into the periplasmic space of the cell. After a purification procedure including ion-exchange chromatography and gel filtration, about 10mg of homogenous recombinant PA-D4 was obtained from 1 liter culture. Data from N-terminal sequencing suggested that the amino acid sequence of recombinant PA-D4 was identical with its natural counterpart. And the result of Western blot showed that the recombinant protein could bind to anti-PA serum from rabbit. Conclusion High-level expression of PA-D4 was obtained in E. coli. The results reported here are parts of a series research to evaluate PA-D4 as a potential medicine for anthrax therapy or a candidate of new vaccine.