FIG. 2.

Expression and purification of recombinant LcrV and its variants. (A) Full-length wild-type Y. pestis strain KIM LcrV was expressed as an N-terminal decahistidyl fusion protein from the T7 polymerase expression vector pET16b. Recombinant lcrV variant genes encoding staggered 30-amino-acid deletions were generated from PCR-amplified DNA fragments and expressed under the same conditions. (B) rLcrV and rV1 to -11 were purified by affinity chromatography on Ni-NTA and eluted with imidazole. After Triton X-114 extraction of endotoxin, proteins were purified on Sephadex G25. Protein aliquots were separated by SDS-PAGE and stained with Coomassie brilliant blue. M indicates molecular size markers, with mass assignments in kDa. rLcrV was loaded in lane 1, while lanes 2 to 12 harbor rV1 to rV11, in numerical order.