Ectopic expression of BmeryCA in Bombyx mori increases silk yield and mechanical properties by altering the pH of posterior silk gland

• Published in: International journal of biological macromolecules 2024-06, Vol.271 (Pt 2), p.132695, Article 132695
• Main Authors: Shi, Run, Lu, Wei, Yang, Jie, Ma, Sanyuan, Wang, Aoming, Sun, Le, Xia, Qingyou, Zhao, Ping
• Format: Article
• Language: English
• Subjects: acid-base balance; active sites; Bombyx mori; carbon dioxide; carbonate dehydratase; Carbonic anhydrase; class; erythrocytes; genetically modified organisms; hemolymph; Mechanical properties; mutants; protein synthesis; secretion; silk; silk glands; silk proteins; Silk yield
• ISSN: 0141-8130; 1879-0003; 1879-0003
• DOI: 10.1016/j.ijbiomac.2024.132695

The silk glands are the specialized tissue where silk protein synthesis, secretion, and conformational transitions take place, with pH playing a critical role in both silk protein synthesis and fiber formation. In the present study, we have identified erythrocyte carbonic anhydrase (BmeryCA) belonging to the α-CA class in the silk gland, which is a Zn2+ dependent metalloenzyme capable of efficiently and reversibly catalyzing the hydrated reaction of CO2 to HCO3−, thus participating in the regulation of acid-base balance. Multiple sequence alignments revealed that the active site of BmeryCA was highly conserved. Tissue expression profiling showed that BmeryCA had relatively high expression levels in hemolymph and epidermis but is barely expressed in the posterior silk gland (PSG). By specifically overexpressing BmeryCA in the PSG, we generated transgenic silkworms. Ion-selective microelectrode (ISM) measurements demonstrated that specifically overexpression of BmeryCA in the PSG led to a shift in pH from weakly alkaline to slightly neutral conditions. Moreover, the resultant PSG-specific BmeryCA overexpression mutant strain displayed a significant increase in both silk yield and silk fiber mechanical properties. Our research provided new insights into enhancing silk yield and improving the mechanical properties of silk fibers. •The active sites and zinc ion binding sites of BmeryCA were both highly conserved.•Overexpression of BmeryCA in the PSG caused the pH of the PSG to change from weakly alkaline to slightly neutral.•Overexpression of BmeryCA in the PSG resulted in increased silk yield and improved mechanical properties of the silk fibers.