Acid protease activity of a major surface membrane glycoprotein (gp63) from Leishmania mexicana promastigotes.
A unique protease with activity optimal at pH 4.0 and trailing toward the alkaline pH spectrum was detected with intact glutaraldehyde-fixed promastigotes of Leishmania mexicana amazonensis, indicating surface localisation of the enzyme. That this surface protease may be a virulence factor is suggested by its apparent roles in multiple steps during leishmanial infections of macrophages. Indeed, its specific activity was 2-2.5 fold higher on virulent cells than on avirulent cells. Several lines of evidence indicate that this acid protease activity is expressed by the major surface glycoprotein (gp63) of L. m. amazonensis. Monoclonal antibody affinity purified gp63 degraded serum albumin, hemoglobin, complement C3, immunoglobulin G and purified rat liver lysosomal proteins in their native forms. The specific activity is about 20-fold higher at pH 4.0 than at pH 7.5 and is about four-fold higher at the body temperature of the mammalian host (37 degrees C) than at that of the insect host (27 degrees C). The protease activity is sodium dodecyl sulphate-sensitive. Among various protease inhibitors tested, only heavy metal ions (1 mM), 1,10-orthophenanthroline (1 mM) and bestatin (100 ng ml-1) significantly inhibited gp63 acid protease activity by up to 80%. N-linked oligosaccharides of gp63 appear to be important for the stability of this molecule, possibly by preventing its autodegradation. Purified gp63 effected limited proteolysis of human complement C3 molecules at the physiological serum pH of 7.5 in a manner, which supports the idea of its participation in complement-receptor mediated endocytosis of promastigotes by macrophages.
Chaudhuri
G
G
Department of Microbiology, University of Health Sciences/The Chicago Medical School, IL 60064.
Chang
K P
KP
D000818:Animals; D000953:Antigens, Protozoan / Q000378:metabolism*; D016282:Aspartic Acid Endopeptidases; D003176:Complement C3 / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism*; D006454:Hemoglobins / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000378:metabolism; D007894:Leishmania mexicana / Q000201:enzymology* / Q000472:pathogenicity; D008247:Lysosomes; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008666:Metalloendopeptidases; D011506:Proteins / Q000378:metabolism; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013696:Temperature; D014774:Virulence
D000953:Antigens, Protozoan; D003176:Complement C3; D006454:Hemoglobins; D007074:Immunoglobulin G; D008562:Membrane Glycoproteins; D011506:Proteins; C018589:lysosomal proteins; D012710:Serum Albumin, Bovine; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; C055146:glycoprotein gp63, Leishmania