Carboxy-terminal conversion of profibrillin to fibrillin at a basic site by PACE/furin-like activity required for incorporation in the matrix

Fibrillin-1, the main component of 10-12 nm microfibrils of the extracellul ar matrix, is synthesized as profibrillin and proteolytically processed to fibrillin, The putative cleavage site has been mapped to the carboxy-termin al domain of profibrillin-1, between amino acids arginine 2731 and serine 2 732, by a spontaneous mutation in this recognition site that prevents profi birillin conversion. This site contains a basic amino acid recognition sequ ence (R-C-R-K-R-R) for proprotein convertases of the furin/PACE family, In this study, we use a mini-profibrillin protein to confirm the cleavage in t he carboxy-terminal domain by both fibroblasts and recombinantly expressed furin/PACE, PACE4, PC1/3 and PC2, Site-directed mutagenesis of amino acids in the consensus recognition motif prevented conversion, thereby identifyin g the scissile bond and characterizing the basic amino acids required for c leavage. Using a PACE/furin inhibitor, we show that wild-type profibrillin is not incorporated into the extracellular matrix until it is converted to fibrillin, Therefore, profibrillin-1 is the first extracellular matrix prot ein to be shown to be a substrate for subtilisin-like proteases, and the co nversion of profibrillin to fibrillin controls microfibrillogenesis through exclusion of uncleaved profibrillin.