IgA1 PROTEASES

Abstract

IgA1 protease is elaborated by several mucosal pathogens, including Neisseria gonorrhoeae, Neisseria meningitidis, Haemophilus influenzae and Streptococcus pneumoniae, as well as Streptococcus sanguis, a predominant bacterium in dental plaque. The enzyme is specific for the IgA1 subclass, cleaving the molecule at the hinge region to form intact F ab and F c fragments. IgA2 is not susceptible to proteolysis because of a deletion of a 13 amino acid segment in the hinge region where the cleavage sites are located. S. sanguis 10556 was grown in carbon excess and carbon limitation, in undialyzed and diafiltrated media, and the enzyme partially purified by gel filtration and anion exchange chromatography using high performance liquid chromatography (HPLC). Comparison was made of the elution profiles of the enzyme preparations in gel filtration using phosphate buffer, pH 7.0, and Tris(hydroxymethyl)amino methane-HCL (Tris-HCL) buffer, pH 8.0. IgA1 protease was found to be elaborated under carbon limitation, a condition encountered by microorganisms in dental plaque. The use of diafiltrated medium reduced significantly a large contaminating peak which elutes in approximately the same region as the enzyme in gel filtration (phosphate buffer). Better separation of the enzyme from other components eluting at the void volumn was obtained using phosphate buffer than Tris-HCL buffer in gel filtration. In anionic exchange chromatography, the enzyme eluted unbound, while the majority of the contaminating proteins remained bound to the support. This procedure is an efficient method of obtaining a preparation of S. sanguis IgA1 protease which is contaminated by only a few proteins.