Characterization of wheat puroindoline proteins.
Puroindoline proteins were purified from selected UK-grown hexaploid wheats. Their identities were confirmed on the basis of capillary electrophoresis mobilities, relative molecular mass and N-terminal amino acid sequencing. Only one form of puroindoline-a protein was found in those varieties, regardless of endosperm texture. Three allelic forms of puroindoline-b protein were identified. Nucleotide sequencing of cDNA produced by RT-PCR of isolated mRNA indicated that these were the 'wild-type', found in soft wheats, puroindoline-b containing a Gly-->Ser amino acid substitution (position 46) and puroindoline-b containing a Trp-->Arg substitution (position 44). The latter two were found in hard wheats. Microheterogeneity, due to short extensions and/or truncations at the N-terminus and C-terminus, was detected for both puroindoline-a and puroindoline-b. The type of microheterogeneity observed was more consistent for puroindoline-a than for puroindoline-b, and may arise through slightly different post-translational processing pathways. A puroindoline-b allele corresponding to a Leu-->Pro substitution (position 60) was identified from the cDNA sequence of the hard variety Chablis, but no mature puroindoline-b protein was found in this or two other European varieties known to possess this puroindoline-b allele. Wheats possessing the puroindoline-b proteins with point mutations appeared to contain lower amounts of puroindoline protein. Such wheats have a hard endosperm texture, as do wheats from which puroindoline-a or puroindoline-b are absent. Our results suggest that point mutations in puroindoline-b genes may confer hard endosperm texture through accumulation of allelic forms of puroindoline-b proteins with altered functional properties and/or through lower amounts of puroindoline proteins.
Day
Li
L
The University of Reading, School of Food Biosciences, Reading, UK. li.day@csiro.au
Bhandari
Dhan G
DG
Greenwell
Philip
P
Leonard
Steven A
SA
Schofield
J David
JD
D000483:Alleles; D002266:Carboxymethylcellulose Sodium / Q000737:chemistry / Q000378:metabolism; D019075:Electrophoresis, Capillary; D013058:Mass Spectrometry; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D017354:Point Mutation; D014908:Triticum / Q000737:chemistry / Q000235:genetics*
D010940:Plant Proteins; C082942:puroindoline protein, Triticum aestivum; D002266:Carboxymethylcellulose Sodium