The survival and replication of Brucella in macrophages is initially triggered by a low intraphagosomal pH. In order to identify
proteins released by Brucella during this early acidification step, we analyzed Brucella suis
conditioned medium at various pH levels. No significant
proteins were released at pH 4.0 in minimal medium or
citrate buffer, whereas in
acetate buffer, B. suis released a substantial amount of soluble
proteins. Comparison of 13 N-terminal amino acid sequences determined by Edman degradation with their corresponding genomic sequences revealed that all of these
proteins possessed a
signal peptide indicative of their periplasmic location. Ten
proteins are putative substrate
binding proteins, including a homologue of the
nopaline binding protein of Agrobacterium tumefaciens. The absence of this homologue in Brucella melitensis was due to the deletion of a 7.7-kb
DNA fragment in its genome. We also characterized for the first time a hypothetical 9.8-kDa basic
protein composed of five
amino acid repeats. In B. suis, this
protein contained 9 repeats, while 12 were present in the B. melitensis orthologue. B. suis in
acetate buffer depended on neither the virB type IV secretory system nor the omp31 gene product. However, the integrity of the omp25 gene was required for release at acidic pH, while the absence of omp25b or omp25c displayed smaller effects. Together, these results suggest that Omp25 is involved in the membrane permeability of Brucella in acidic medium.