Influence of variation in high-molecular-weight glutenin subunits composition at Glu-B1 and Glu-D1 loci on gluten microstructure and northern Chinese steamed bread quality

1. Near-isogenic and chromosome substitution lines with uniform genetic backgrounds were employed to systematically reveal the effects of gluten proteins on gluten structure, flour properties, and northern Chinese steamed bread (NCSB) quality. 2. A comprehensive evaluation of wheat processing quality was achieved by integrating multiple analytical techniques, highlighting the advantages of confocal laser scanning microscopy (CLSM) and laser confocal microscopy Raman (LCM-Raman). 3. Thermogravimetric analysis (TGA) revealed weight loss patterns at 150°C and 600°C that closely correlated with steamed bread quality and overall bread quality, respectively. 4. Superior steamed bread quality is not necessarily dependent on higher gluten strength. High-molecular-weight glutenin subunits (HMW-GS) play a key role in determining wheat processing quality, but their contribution to NCSB quality has not been systematically elucidated. In this study, near-isogenic lines and chromosome substitution lines differing in HMW-GS compositions at the Glu-B1 and Glu-D1 loci were utilized to comprehensively assess the impact of gluten proteins on gluten microstructure, flour functionality, and NCSB quality through multiple analytical methods. Unextractable polymeric protein content (%UPP), disulfide bond content, TGA, rapid viscosity analyzer (RVA) profiles, gluten aggregation behavior, and steamed bread-making trials were used to evaluate gluten structure and functional performance. CLSM and LCM-Raman analyses demonstrated that CB037B and SL/B significantly increased junction density, protein network area, and β-sheet content, while reducing lacunarity and α-helix content. Steamed bread-making trials combined with dough rheological assessments revealed that although CB037B exhibited strong-gluten characteristics, the resulting steamed bread quality was lower than that of CB037A, supporting the concept that superior NCSB quality does not necessarily depend on higher gluten strength. TGA further indicated that the weight loss pattern at 150°C (CB037A>CB037B>SL/B>CS) aligned with steamed bread quality rankings, whereas weight loss at 600°C corresponded more closely with conventional bread quality. Overall, this work provides valuable insights into the structure–function relationships of HMW-GS in wheat and introduces novel indicators for predicting both steamed bread and bread quality, offering guidance for breeding programs aimed at improving wheat processing performance in northern China.