Inhibitory serpins from wheat grain with reactive centers resembling glutamine-rich repeats of prolamin storage proteins - Cloning and characterization of five major molecular forms

被引:107
作者
Ostergaard, H
Rasmussen, SK
Roberts, TH
Hejgaard, J
机构
[1] Tech Univ Denmark, Dept Biochem & Nutr, DK-2800 Lyngby, Denmark
[2] Riso Natl Lab, Plant Biol & Biogeochem Dept, DK-4000 Roskilde, Denmark
关键词
D O I
10.1074/jbc.M004633200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Genes encoding proteins of the serpin superfamily are widespread in the plant kingdom, but the properties of very few plant serpins have been studied, and physiological functions have not been elucidated. Six distinct set-pins have been identified in grains of hexaploid bread wheat (Triticum aestivum L.) by partial purification and amino acid sequencing. The reactive centers of all but one of the serpins resemble the glutamine-rich repetitive sequences in prolamin storage proteins of wheat grain. Five of the serpins, classified into two protein Z subfamilies, WSZ1 and WSZ2, have been cloned, expressed in Escherichia coli, and purified. Inhibitory specificity toward 17 proteinases of mammalian, plant, and microbial origin was studied. All five serpins were suicide substrate inhibitors of chymotrypsin and cathepsin G. WSZ1a and WSZ1b inhibited at the unusual reactive center P-1-P-1' Gln-Gln, and WSZ2b at P-2-P-1 Leu-Arg-one of two overlapping reactive centers. WSZ1c with P-1-P-1' Leu-Gln was the fastest inhibitor of chymotrypsin (k(a) = 1.3 x 10(6) M-1 s(-1)). WSZ1a was as efficient an inhibitor of chymotrypsin as WSZ2a (k(a) similar to 10(5) M-1 s(-1)), which has P-1-P-1' Leu-Ser-a reactive center common in animal serpins. WSZ2b inhibited plasmin at P-1-P-1' Arg-Gln (k(a) similar to 10(3) M-1 s(-1)). None of the five serpins inhibited Bacillus subtilisin A, Fusarium trypsin, or two subtilisin-like plant serine proteinases, hordolisin from barley green malt and cucumisin D from honeydew melon. Possible functions involving interactions with endogenous or exogenous proteinases adapted to prolamin degradation are discussed.
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页码:33272 / 33279
页数:8
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