| A substantial portion of the lower molecular weight polypeptides in the glutenin polymer wereα-andγ-gliadins. Theα- andγ-gliadins contain an odd number of cysteine residues were reported. The gliadins with an odd number of cysteines could form intermolecular disulfide bond and thus participate in the gluten polymer. Changes in position of cysteine residues might affect the pattern of disulfide bond formation, resulting in a failure of two cysteine residues in a protein. Two such cysteine residues would then be available for intermolecular disulfide bond formation. The glutamine ofω-gliadin may play an important role in the unique viscoelastic properties of the gluten protein complexes through strong hydrogen bond to interact cohesively with other dough proteins. Compared toα-andγ-gliadins, the number ofω-gliadin gene sequences in Genbank is much smaller. Therefore, it is necessary to further understand the primary structure ofω-gliadins. In this study, the gliadin composition of Chuannong 16 was analyzed by using acid polyacrylamide gel electrophoresis (A-PAGE). Using a PCR-based strategy we isolated and sequenced the gliadin sequences from the wheat varieties Chuannong 16. The main results were as following:1. Using PCR method,5α-gliadin genes, designated as Gli-α-1 to Gli-α-5, were obtained from Chuannong 16. Gli-α-1 and Gli-α-5 are putatively functional, while Gli-α-2, Gli-w-3, Gli-w-4 are recognized as pseudogenes, since the transition from C to T resulting in the in-frame stop codon. Gli-α-3, Gli-α-4 and Gli-α-5 show a typical structure ofα-gliadins:a 20-residue signal peptide, a repetitive domain composed of basic repeating units, two polyglutamine stretches, and two unique sequences which contain six cysteine. Most of the in-frame stop codons instead of glutamine occurred in the polyglutamine stretches. Gli-a-1, Gli-a-2 got a deletion of a unique domain I where four cysteine exist and a polyglutamine region, contain only two cysteine, could form intermolecular disulfide bond and thus participate in the gluten polymer.2. Acid polyacrylamide gel electrophoresis (A-PAGE) results showed that the Chuannong 16 had 7 bands in theωzone. Using PCR method,7ω-gliadin genes, designated as Gli-w-1 to Gli-w-7, were obtained from Chuannong 16. Three of the 7 genes are putatively functional, while Gli-w-1, Gli-w-2, Gli-w-6 and Gli-w-7 are recognized as pseudogenes, since there are frameshift mutations in Gli-w-1 and Gli-w-6, and both Gli-w-2 and Gli-w-7 contain an in-frame stop codon. The deduced protein sequences of the 7 genes show a typical structure of co-gliadins:a 19-residue signal peptide, followed by a 10-11-residue non-repetitive N-terminus, a repetitive region rich in glutamine and proline and a 10-11 residue C-terminus. Alignment of the deduced amino acid sequences indicates that the N-and C-terminal of co-gliadins are relatively conserved, and that the repetitive regions contain much more variations. In terms of the deduced amino acid sequence, all of them belong to ARQ-/ARE type which is also called as co-2 gliadin. Percent of Q (glutamine) is 35-40mol% while Q:P:F is approximately 4:3:1. Analysis showed that the gene base transition from C to T got a high frequency. It caused the two in-frame stop codon of Gli-w-2 and Gli-w-7.It is also responsible for the amino acid replacement from prolinamide (P) to leucine (L), from proline (P) to Ser (S) which appears multiple times in the repeat region of co-gliadin. |
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