PROTEOLYTIC AND NONPROTEOLYTIC ACTIVATION OF HUMAN NEUTROPHIL PROGELATINASE-B

The activation of human neutrophil progelatinase B (pro-HNG) by a vari ety of proteolytic and non-proteolytic activators has been investigate d. A quantitative comparison of the activation efficiencies of treatme nts previously reported to activate pro-HNG or the related gelatinase B species produced by other cells demonstrates that stromelysin and tr ypsin are good activators. HgCl2 is a moderately effective activator, while p-chloromercuribenzoate and NaOCl are poor activators. It is als o shown that human matrilysin and human fibroblast-type collagenase ca n activate pro-HNG by a mechanism that is very similar to that of stro melysin. Initially, these proteinases hydrolyze the Glu(40)-Met(41) bo nd in the propeptide domain to generate an 88 kDa inactive HNG species . Collagenase also generates a 68 kDa HNG species through hydrolysis o f the Ala(74)-Met(75) bond. Ultimately, treatment with either matrilys in, collagenase or trypsin results in the production of a 65 kDa activ e form of HNG that arises from hydrolysis of the Ag-87-Phe(88) bond, T his is the same active species produced on activation by stromelysin. This cleavage site is downstream of the 'cysteine-switch' residue loca ted at position 80 and releases it, accounting for the permanent activ ation of the enzyme. These results suggest that matrilysin and collage nase may be physiologically relevant activators of pro-HNG and/or othe r progelatinase B species. Activation by HgCl2 produces an active 68 k Da enzyme due to autolytic hydrolysis of the Ala(74)-Met(75) bond. Thi s species retains the cysteine switch residue; however, it is shown th at it is only active in the continued presence of HgCl2. Removal of th e HgCl2 restores latency, indicating that this species is reversibly a ctivated by HgCl2, which functions by complexing the sulfhydryl group of the cysteine switch residue and keeping it dissociated from the act ive site zinc atom. Thus, in spite of reports to the contrary, the cys teine switch mechanism can account for the latency and activation of p ro-HNG.