Degradation of gonococcal outer membrane proteins by human neutrophil lysosomal proteases.

Interest in the molecular mechanisms of leukocyte bactericidal activity led us to study the effects of human neutrophil lysosomal proteases on the outer membrane (OM) proteins of Neisseria gonorrhoeae. A protease fraction containing cathepsin G and elastase activity was partially purified by gel filtration chromatography of acetate extracts of purified neutrophil granules. OM was obtained from gonococci by French press-Sarkosyl or by LiCl2 extraction. The principal (protein I) and opacity-associated (proteins II) OM proteins of N. gonorrhoeae were hydrolyzed by lysosomal proteases; proteins II were more susceptible to hydrolysis than protein I. Treatment of whole gonococci, with subsequent purification of OM, or direct treatment of purified OM led to identical hydrolysis of OM proteins by lysosomal proteases as indicated by sodium dodecyl sulfate-polyacrylamide gel patterns. Similarly, hydrolysis of purified OM proteins was identical whether OM was treated with unfractionated granule extract or with the partially purified lysosomal proteases, indicating that the observed hydrolysis by unfractionated lysosomal contents was due solely to the lysosomal protease fraction. Hydrolysis of OM proteins was dependent upon the concentration of proteases, time, and temperature. Hydrolysis of proteins II was observed with as little as 1 microgram of proteases per ml for 1 h at 37 degrees C. OM incubated alone or with heat-inactivated proteases showed no hydrolytic activity. The addition of 25 mM Na+, K+, Mg2+, or Ca2+ to incubation mixtures containing proteases and OM did not alter hydrolytic activity as indicated by sodium dodecyl sulfate-polyacrylamide gel patterns.