FIG. 5.

(A) The secA2(K115R) allele does not complement the M. tuberculosis secA2 mutant phenotype in macrophages. Murine bone marrow-derived macrophages were infected at a multiplicity of infection of 1.0 with the strain H37Rv, the ΔsecA2 mutant, the ΔsecA2 mutant complemented by the addition of WT secA2, (ΔsecA2+secA2), and the ΔsecA2 mutant with secA2(K115R) (ΔsecA2+secA2 KR). CFU were determined by plating macrophage lysates at various times postinfection. The infection was performed with triplicate wells for each strain per time point, and the error bars represent means ± standard deviations for the triplicate wells. The symbols for H37Rv and the complemented strain are overlapping at most time points in the graph presented. Data are representative of three independent experiments. *, data are statistically significantly different (P < 0.05). (B) Expression of SecA1 and SecA2 in strains with different secA2 alleles. Equal amounts of formalin-fixed whole-cell lysates of the M. tuberculosis H37Rv strain carrying an empty vector, the M. tuberculosis ΔsecA2 mutant with an empty vector, the ΔsecA2 mutant with the WT secA2 integrated at the chromosomal attB site (WT), and the ΔsecA2 mutant with the secA2(K115R) allele integrated at the attB site (KR) were run on SDS-polyacrylamide gels and subjected to Western blot analysis with anti-SecA1 or anti-SecA2 antibodies. The top panel shows SecA1 protein and the lower panel shows SecA2 protein. MW, molecular weight (in thousands).