HEMAGGLUTINATING ACTIVITY OF EXTRACELLULAR ALKALINE METALLOENDOPEPTIDASES FROM VIBRIO SP. NUF-BPP1

Alkaline metalloendopeptidase (metalloprotease) AP1 (48 kDa) from Vibr io sp. isolated from the intestine of a five-barred goatfish (Parupene us trifasciatus) was reported in our previous paper to produce AP2 (36 kDa) by releasing a peptide fragment (molecular mass of about 12 kDa) from the C-terminal end of AP1 by autodigestion.(1)) AP1 strongly agg lutinated fish (flounder, Paralichthys olivaceus) and rabbit erythrocy tes, and weakly chicken erythrocytes. In contrast, AP2 had no signific ant hemagglutinating activity toward any erythrocytes tested, except f or weak activity on flounder erythrocytes, suggesting that the C-termi nal region of AP1 may be required for the strong hemagglutinating acti vity. The optimum temperature for the hemagglutinating activity of AP1 was found to be lower than that for the proteolytic activity. At acid ic pHs (below pH 7.5), the hemagglutinating activity of AP1 decreased, and its pH profile resembled that of the proteolytic activity. The he magglutinating activity of AP1 was not observed in the presence of o-p henanthroline or synthetic and proteinous substrates, but different ki nds of saccharides and lipids had no effect. While the proteolytic act ivity of AP1 was not affected by CaCl2, the hemagglutinating activity of AP1 decreased with increases in CaCl2 concentrations. These results suggested that the hemagglutinating activity of these proteases (AP1 and AP2) was most likely caused by their proteolytic action on erythro cyte cell surfaces.