The role of L-valine in synthetic biology

L-valine plays an important role in synthetic biology, exerting key functions in multiple links from metabolic pathway reconstruction to product synthesis. The details are as follows:

Metabolic Pathway Reconstruction

Relieving feedback inhibition: Acetohydroxyacid synthase (AHAS), a key enzyme in the L-valine synthesis pathway, is susceptible to feedback inhibition by L-valine and other branched-chain amino acids. Site-directed mutagenesis of the regulatory subunit of AHAS, such as mutating specific amino acid sites to Asp-Asp-Phe, can relieve this feedback inhibition, maintain enzyme activity, and thus allow metabolic flux to flow more smoothly toward L-valine synthesis, increasing its yield.

Optimizing carbon metabolic flux: Pyruvate is the main precursor for L-valine synthesis. By knocking out or modifying genes related to pyruvate-competing metabolism, such as lactate dehydrogenase gene (ldhA) and pyruvate quinone oxidoreductase gene (pqo), the diversion of pyruvate to other pathways (e.g., lactate, acetic acid) can be reduced, and the carbon flux of pyruvate toward L-valine synthesis pathway can be increased. Meanwhile, overexpressing key enzyme genes in the glycolytic pathway, such as fructose-6-phosphate kinase encoding gene (pfkA) and pyruvate kinase encoding gene (pyk), can promote glucose metabolism, generate more pyruvate, and provide sufficient precursors for L-valine synthesis.

Balancing cofactor supply: Cofactors are required in the L-valine synthesis process. For example, the reaction catalyzed by acetohydroxyacid isomeroreductase requires NADPH as a coenzyme. By optimizing metabolic pathways and regulating the expression of related enzymes, the balance of intracellular cofactors can be maintained, providing sufficient coenzymes for L-valine synthesis and ensuring the smooth progress of the reaction.

Product Synthesis

Efficient synthesis as a target product: Using synthetic biology technologies, with chassis cells such as Corynebacterium glutamicum and Escherichia coli, efficient synthesis of L-valine can be achieved through metabolic pathway reconstruction. For instance, genetic engineering modification of Corynebacterium glutamicum by overexpressing key genes in the L-valine synthesis pathway (e.g., ilvBN, ilvC, ilvD, ilvE), while enhancing the expression of transporter encoding gene brnFE and its regulatory protein encoding gene lrp1, the resulting engineered strains can mass-produce L-valine in fermenters.

Participating in the synthesis of complex biomolecules: As one of the basic units of proteins, L-valine can be used as a raw material in synthetic biology to participate in the synthesis of functional proteins or peptide drugs. By precisely controlling amino acid sequences and introducing L-valine into peptide chains in a specific order, biologically active polypeptides (such as certain antimicrobial peptides and hormone analogs) can be synthesized, laying a foundation for drug development.

Used in constructing cell factories: With the goal of efficient L-valine synthesis, overall modification of cells can build high-efficiency cell factories. Through multi-module engineering strategies, not only are its synthesis pathways and transport modules optimized, but transcription factors related to its synthesis are also regulated. For example, overexpressing pdhR and inhibiting the expression of rpoS can promote L-valine synthesis; meanwhile, introducing heterologous pathways to enhance NADPH supply can improve the efficiency and yield of L-valine production in cell factories.