The role of asparagine-linked glycosylation site on the catalytic domain of matriptase in its zymogen activation.
Matriptase is a type II transmembrane serine protease containing one potential site for asparagine-linked glycosylation (N-glycosylation) on the catalytic domain (Asn772). It has been found that the activation of matriptase zymogen occurs via a mechanism requiring its own activity and that the N-glycosylation site is critical for the activation. The present study aimed to determine the underlying reasons for the site requirement using Madin-Darby canine kidney cells stably expressing recombinant variants of rat matriptase. A full-length variant with glutamine substitution at Asn772 appeared to be unable to undergo activation because of its catalytic incompetence (i.e., decreased availability of the soluble catalytic domain and/or of the correctly folded domain). This was evidenced by the observations that (i) a recombinant catalytic domain of matriptase with glutamine substitution at the site corresponding to matriptase Asn772 [N772Q-CD-Myc(His)(6)] was not detected in the medium conditioned by transfected cells but was on the cell surface and (ii) purified N772Q-CD-Myc(His)(6) exhibited markedly reduced activity toward a peptide substrate. It is concluded that N-glycosylation site at Asn772 of matriptase is required for the zymogen activation because it plays an important role in rendering this protease catalytically competent in the cellular environment.
Miyake
Yuka
Y
Laboratory of Enzyme Chemistry, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
Tsuzuki
Satoshi
S
Mochida
Seiya
S
Fushiki
Tohru
T
Inouye
Kuniyo
K
D000818:Animals; D001216:Asparagine / Q000235:genetics*; D020134:Catalytic Domain / Q000235:genetics; D002460:Cell Line; D004285:Dogs; D004789:Enzyme Activation / Q000502:physiology*; D004792:Enzyme Precursors / Q000737:chemistry*; D006031:Glycosylation; D051381:Rats; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000235:genetics
D004792:Enzyme Precursors; D001216:Asparagine; D012697:Serine Endopeptidases; C485595:matriptase