Immunochemical localization of CAMP factor (protein B) in Streptococcus agalactiae

Biochemical and immunochemical investigations were used in order to study the quantitative and qualitative localization of CAMP factor (protein B) in the cell fractions of Streptococcus agalactiae during the logarithmic growth phase. The dynamic quantitative distribution of CAMP factor activity showed that higher concentrations of CAMP factor were found in the cytoplasm than in the cell envelopes. A maximal intracellular accumulation of CAMP factor activity was observed in the late log phase. Immunoblotting analysis using specific anti-CAMP-IgG showed that CAMP factor could be detected in the different cell fractions of S. agalactiae. During the early log phase, CAMP factor was present as a single 25 kD protein band in the cytoplasm; white it was found together with its 26 and 24 kD satellite proteins in the cytoplasmic membrane and the cell wall as well as in all the cell fractions in the mid- and late log phases. Intracellular CAMP factor exhibited the same antigenic and amphiphilic behaviour as the extracellular species. Additionally, a newly discovered amphiphilic protein of approximately 54 kD which exhibited similar antigenicity with the CAMP factor was present in all the cell fractions. Immunoelectron microscopic examinations using ferritin-labelled antibodies revealed that CAMP factor was mainly found in the cytoplasm, whereas it was associated to a minor extent with the cell envelopes. Interestingly, an accumulation of CAMP factor was also localized either at the sites of cross-wall initiation or at the cell surfaces where the cell wall became autolysed.