Interaction between leukocyte elastase and elastin: quantitative and catalytic analyses

Solubilization of elastin by human leukocyte elastase (HLE) cannot be analy zed by conventional kinetic methods because the biologically relevant subst rate is insoluble and the concentration of enzyme-substrate complex has no physical meaning. We now report quantitative measurements of the binding an d catalytic interaction between HLE and elastin permitted by analogy to rec eptor-ligand systems. Our results indicated that a limited and relatively c onstant number of enzyme binding sites were available on elastin, and that new sites became accessible as catalysis proceeded. The activation energies and solvent deuterium isotope effects were similar for catalysis of elasti n and a soluble peptide substrate by HLE, yet the turnover number for HLE d igestion of elastin was 200-2000-fold lower than that of HLE acting on solu ble peptide substrates. Analysis of the binding of HLE to elastin at 0 degr ees C, in the absence of significant catalytic activity, demonstrated two c lasses of binding sites (K-d = 9.3 X 10(-9) M and 2.5 X 10(-7) M). The high er affinity sites accounted for only 6% of the total HLE binding capacity, but essentially all of the catalytic activity, and dissociation of HLE from these sites was minimal. Our studies suggest that interaction of HLE with elastin in vivo may be very persistent and permit progressive solubilizatio n of this structurally important extracellular matrix component. (C) 1999 E lsevier Science B.V. All rights reserved.