‘Shed’ furin: mapping of the cleavage determinants and identification of its C-terminus

The human endoprotease furin is involved in the proteolytic maturation of the precursor molecules of a wide variety of bioactive proteins. Despite its localization in the membranes of the trans-Golgi system by means of a transmembrane domain, it has repeatedly been reported to form a C-terminally truncated, naturally secreted form referred to as ‘shed’ furin. In order to identify the cleavage site, internal deletion mutants of increasing size, N-terminal to Leu708, and subsequently individual amino acid substitutions were introduced, and Arg683 was identified as the prime determinant for shedding. MS analysis determined Ser682 as the C-terminus of shed furin, suggesting that monobasic cleavage may occur N-terminal to Arg683. Alteration of Arg683 directs the shedding mechanism to alternative cleaving sites previously unused.