Molecular characterisation of the low-molecular weight glutenin subunit genes of tall wheatgrass and functional properties of one clone Ee34.
Wild tall wheatgrass (Lophopyrum elongatum L., 2x = 14) is an important resource for improving bread wheat (Titicum aestivum L.), including HMW-GS and LMW-GS relevant to end-use quality of the wheat flour. A set of 14 distinct sequences were amplified from the genomic DNA of the tall wheatgrass, using degenerate primers targeted at Glu-3, the locus containing the genes encoding the low-molecular weight glutenin subunits (LMW-GS). Three sequences contained an internal stop codon and were classified as pseudogenes. The other 11 all consisted of a single intron-less intact open-reading frame. An alignment of deduced protein sequences showed that the primary structure of all 11 sequences was similar to that of wheat and other wheat-related grass Glu-3 genes. All 11 sequences carried the 14 amino acid residue N-terminal motif MESNIIISFLK/RPWL, and were classified as LMW-m genes, based on the identity of the first amino acid of the mature protein. All but one of the sequences contained seven cysteine residues (the exception had 6). Their repetitive domain differs significantly from that present in Glu-3 genes isolated from the close relative intermediate wheatgrass (Thinopyrum Intermedium, 6x). A phylogenetic analysis showed that the tall wheatgrass sequences were closely related to those of the intermediate wheatgrass, but only distantly so to those from decaploid tall wheatgrass. One of the 11 LMW-GS peptides with a free-cysteine residue was heterologously expressed in E. coli and purified in sufficient scale to perform a flour supplementation test. This showed that the dough strength of bread wheat flour was significantly increased by the presence of the tall wheatgrass LMW-GS.
Chen
Fanguo
F
The Key Laboratory of Plant Cell Engineering and Germplasm Innovation of Ministry of Education, School of Life Science, Shandong University, 250100, Jinan, People's Republic of China.
Liu
Shuwei
S
Zhao
Feng
F
Xu
Chunhui
C
Xia
Guangmin
G
D000595:Amino Acid Sequence; D001939:Bread / Q000032:analysis; D016384:Consensus Sequence; D030562:Databases, Protein; D005433:Flour / Q000032:analysis; D005503:Food Additives / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D005511:Food Handling / Q000379:methods; D017343:Genes, Plant; D005983:Glutens / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D055595:Mechanical Phenomena; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D030821:Plants, Genetically Modified; D006109:Poaceae / Q000737:chemistry / Q000235:genetics*; D021122:Protein Subunits / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D011786:Quality Control; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D014908:Triticum / Q000737:chemistry
D005503:Food Additives; D021122:Protein Subunits; D011994:Recombinant Proteins; D005983:Glutens; C037302:glutenin