By anion-exchange chromatography of the concentrated culture fluids of P. gingivalis W 50, two proteolytic enzymes, referred to as RGP-I and RGP-II were, eluted from column at the different NaCl concentrations in the eluant buffer. The two enzymes were purified until the preparations displayed single bands with sodium dodecyl sulfate-polyacryl amide gel electrophoresis (SDS-PAGE) by gel filtration and isoelectric focusing. Comparative studies between the two enzymes revealed that the same molecular masses (44 kDa) of RGP-I and II were obtained both with gel filtration and SDS-PAGE. However, the isoelectric points of RGP-I and II were 5.0 and 5.7, respectively. Both enzymes were thiol proteinases and were inhibited by sulfhydryl group blocking reagents, tosyl-L-lysine chloromethylketone, EDTA, leupeptin, L-trans-epoxy-succinylleucylamido-(4-guanidino) butane, and antipain. Both enzymes were activated by glycylglycine. RGP-I and RGP-II hydrolyzed synthetic substrates containing arginine in the P-1 positions, but those containing lysine in the same position were notcleaved
