Fetuin-A and Cystatin C Are Endogenous Inhibitors of Human Meprin Metalloproteases

Meprin alpha and beta zinc metalloproteinases, play significant roles in inflammation, including inflammatory bowel disease (IBD), possibly by activating cytokines, like interleukin 1 beta, interleukin 18, or tumor growth factor alpha. Although a number of potential activators for meprins are known, no endogenous inhibitors have been identified. In this work, we analyzed the inhibitory potential of human plasma and identified bovine fetuin-A as an endogenous meprin inhibitor with a K-i (inhibition constant) of 4.2 x 10(-5) M for meprin alpha and a K-i of 1.1 x 10(-6) M meprin beta. This correlated with data obtained for a fetuin-A homologue from carp (nephrosin inhibitor) that revealed a potent meprin alpha and beta inhibition (residual activities of 27 and 22%, respectively) at a carp fetuin concentration of 1.5 x 10(-6) M. Human fetuin-A is a negative acute phase protein involved in inflammatory diseases, thus being a potential physiological regulator of meprin activity. We report kinetic studies of fetuin-A with the proteolytic enzymes astacin, LAST, LAST_MAM, trypsin, and chymotrypsin, indeed demonstrating that fetuin-A is a broad-range protease inhibitor. Fetuin-A inhibition of meprin beta activity was 40 times weaker than that of meprin beta activity. Therefore, we tested cystatin C, a protein structurally closely related to fetuin-A. Indeed, cystatin C was an inhibitor for human meprin alpha (K-i = 8.5 x 10(-6) M) but, interestingly, not for meprin beta. Thus, the identification of fetuin-A and cystatin C as endogenous proteolytic regulators of meprin activity broadens our understanding of the proteolytic network in plasma.