Nephelometric assay for the immunoglobulin A1-protease produced by the oral bacterium Streptococcus sanguis

Streptococcus sanguis, an initiator of human dental plaque, produces an endopeptidase which cleaves immunoglobulin A1 (IgA1) at the hinge region. A rapid nephelometric assay was developed for the quantitation of IgA1-specific protease activity. The protease was produced in dialysis cultures which yielded cell-free fluids having 14 times the specific activity of conventional cultures. Assay was based on the difference in detectable IgA1 concentrations at the start and termination of the reaction; IgA1 concentrations were determined by rate of complex formation with IgA-specific antibody. The rates of IgA1 cleavage were linear during incubations up to 3 h if enzyme preparations were sufficiently diluted. The assay resolution was less than that achieved with sodium dodecyl sulphate-polyacrylamide gel electrophoresis, but 1 h incubation of protease-IgA1 reaction mixtures was adequate for measurement. The pH optimum for the reaction was 7.0 and the calculated Km was 5.6 X 10(-5)M IgA1. The optimal incubation temperature was in the range of 37-40 degrees C; the enzyme lost all activity at 55 degrees C.