Inhibitory serpins from wheat grain with reactive centers resembling glutamine-rich repeats of prolamin storage proteins. Cloning and characterization of five major molecular forms.
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 serpins 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) approximately 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) approximately 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.
Ostergaard
H
H
Department of Biochemistry and Nutrition, Bldg. 224, Technical University of Denmark, DK-2800 Lyngby, Denmark.
Rasmussen
S K
SK
Roberts
T H
TH
Hejgaard
J
J
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D002523:Edible Grain / Q000737:chemistry*; D005973:Glutamine; D006031:Glycosylation; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D055315:Prolamins; D011994:Recombinant Proteins / Q000737:chemistry; D020449:Repetitive Sequences, Amino Acid; D015843:Serpins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D014908:Triticum / Q000737:chemistry*
D010940:Plant Proteins; D055315:Prolamins; D011994:Recombinant Proteins; D015843:Serpins; C099007:WSZ1 protein, Triticum aestivum; D005973:Glutamine