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United States Department of Agriculture

Agricultural Research Service

Title: Atomic Force Microscopy of a Hybrid High-Molecular-Weight Glutenin Subunit from a Transgenic Hexaploid Wheat

Authors
item Mcintire, Theresa - UNIV OF CA, RIVERSIDE
item Lew, Ellen
item Adalsteins, Elva - DECODE GENETICS
item Blechl, Ann
item Anderson, Olin
item Brant, David - UNIV OF CA, RIVERSIDE
item Kasarda, Donald

Submitted to: Biopolymers
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: December 16, 2004
Publication Date: January 1, 2005
Citation: McIntire, T.M., Lew, E.J.L., Adalsteins, A.E., Blechl, A., Anderson, O.D., Brant, D.A., Kasarda, D.D. 2005. Atomic force microscopy of a hybrid high-molecular-weight glutenin subunit from a transgenic hexaploid wheat. Biopolymers. 78(2):53-61.

Interpretive Summary: The high-molecular-weight glutenin subunits (HMW-GS) show the strongest correlations with variations in wheat quality characteristics of all the wheat grain protein types. The molecular basis for these correlations has not been clearly established, but the large repeating sequence domain of the HMW-GS is likely to contribute in an important way. We have examined the structure of this domain by way of non-contact mode atomic force microscopy. Our results suggest that the previously held model, the rigid beta-spiral structure, is probably incorrect and the a polyproline II (PPII) helix is more likely. The PPII helix representing a more extended polypeptide chain should favor more extensive side-by-side chain interactions through hydrogen bonding. Improved dough elasticity should result from an enhancement of repeating sequence domain length.

Technical Abstract: The high-molecular-weight glutenin subunits (HMW-GS) of wheat gluten in their native form are incorporated into an intermolecularly disulfide-linked, polymeric system that gives rise to the elasticity of wheat flour doughs. These protein subunits range in molecular weight from about ''K-90K and are made up of small N-terminal and C-terminal domains and a large central domain that consists of repeating sequences rich in glutamine, proline, and glycine. The cysteines involved in forming intra- and intermolecular disulfide bonds are found in, or close to, the N- and C-terminal domains. A model has been proposed in which the repeating sequence domain of the HMW-GS forms a rod-like b-spiral with length near 50 nm and diameter near 2 nm. We have sought to examine this model by using noncontact atomic force microscopy (NCAFM) to image a hybrid HMW-GS in which the N-terminal domain of subunit Dy10 has replaced the N-terminal domain of subunit Dx5. This hybrid subunit, coded by a transgene overexpressed in transgenic wheat, has the unusual characteristic of forming, in vivo, not only polymeric forms, but also a monomer in which a single disulfide bond links the C-terminal domain to the N-terminal domain, replacing the two intermolecular disulfide bonds normally formed by the corresponding cysteine side chains. No such monomeric subunits have been observed in normal wheat lines, only polymeric forms. NCAFM of the native, unreduced 93K monomer showed fibrils of varying lengths but a length of about 110 nm was particularly noticeable whereas the reduced form showed rod-like structures with a length of about 300 nm or greater. The 110 nm fibrils may represent the length of the disulfide linked monomer, in which case they would not be in accord with the b-spiral model, but would favor a more extended conformation for the polypeptide chain, possibly polyproline II.

Last Modified: 10/25/2014
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