Molecular characterisation of high molecular weight glutenin allele Glu-B1h encoding 1Bx14+1By15 subunits in bread wheat (Triticum aestivum L.)

Abstract

In this study, the authentic high molecular weight glutenin (HMW-GS) allele Glu-B1 h encoding for subunits 1Bx14 and 1By15 from German bread wheat cultivars Hanno and Imbros was identified and cross-verified by a suite of established protein analysis technologies, including sodium dodecyl sulfate-polyacrylamide gel electrophoresis, reversed-phase high-performance liquid chromatography, reversed-phase ultra-performance liquid chromatography, and matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF-MS). The complete encoding sequences were isolated by allele-specific PCR, and consist of 2367 bp for 1Bx14 and 2151 bp for 1By15 and encode 789 and 717 amino acid residues, respectively. The deduced molecular masses of two subunit genes were 82 340.13 Da and 74 736.13 Da, corresponding well to those determined by MALDI-TOF-MS. The presence and authenticity of 1Bx14 and 1By15 subunits were further confirmed by liquid chromatography coupled to tandem mass spectrometry and heterologous expression in E. coli. Comparative analysis demonstrated that 1Bx14 possessed one deletion and 20 single-nucleotide polymorphism variations compared with seven other Glu-B1 x-type HMW-GS genes that mainly resulted from C–T substitutions, whereas compared with five other Glu-B1 y-type HMW-GS genes, 1By15 displayed few variations. Phylogenetic analysis based on the complete coding sequences of the published HMW-GS genes showed that 1Bx14 had a high divergence with other 1Bx subunit genes, whereas 1By15 displayed greater similarity with 1By20. A possible evolutionary route for 1Bx14 gene formation is proposed, which might have resulted from an intra-strand illegitimate recombination event that occurred ~1.32 million years ago.

Low molecular weight glutenin subunit gene Glu-B3h confers superior dough strength and breadmaking quality in wheat (Triticum aestivum L.)

“…Our results showed that Glu-B3h contains 1179 bp with a large insert fragment at nucleotides 464–505 that represent 14 amino acids ( Table 2 ), which is longer than most other LMW-GS genes. In addition, the repetitive domain of Glu-B3h contains 15 hexapeptides (consensus PPFSQQ and QQPVLP) and 2 nonapeptides (QQPSFSQQQ), which account for 57% of the repetitive domain that is mainly responsible for the length of LMW-GS and contributes to superior dough quality 34 . These structural features may thus contribute to superior gluten quality.…”

Detection of cysteine residue numbers in wheat gluten proteins by MALDI-TOF

“…For example, a protein with 4 cysteine residues will increase its Mr by 420.56 Da. The x-type HMW-GS usually have 2, 4, or 5 cysteine residues, while most y-type subunit have 6 or 7 cysteine residues [8,27,28].…”

Isolation and molecular characterization of two novel HMW-GS genes from Chinese wheat (Triticum aestivum L.) landrace Banjiemang

“…The subunits 1Ax2*, 1Dx5?1Dy10, 1Bx17?1By18 are positively associated with bread-making qualities in contrast with null, 1Dx2?1Dy12, 1Bx7?1By8 (Branlard and Dardevet 1985;Payne 1987;Rogers et al 1989). The characterization of such quality related to HMW subunits in bread wheat leads to searching for new alleles of HMW-GS in common wheat and its related species (Wan et al 2002;Wang et al 2004;Cong et al 2007;Pang and Zhang 2008;Guo et al 2010;Jiang et al 2005Jiang et al , 2010Jin et al 2012;Ma et al 2013;Xiao et al 2014). Considerable numbers of studies showed that alleles of HMW-GS at one individual locus are rich in Chinese wheat landraces.…”