Research advances in biosynthesis of trans-4-hydroxy-L-proline

The trans-4-hydroxy-L-proline (i.e., T-4-Hyp), a key component of collagen, possesses broad application value in pharmaceuticals, food, cosmetics, and chemical industries. Traditional methods such as animal tissue extraction and chemical synthesis are constrained by low efficiency, significant environmental pollution, and high costs. In contrast, microbial fermentation has emerged as a research focus for the industrial production of T-4-Hyp due to its environmental compatibility and sustainable nature. This review systematically summarizes recent advances in the biosynthesis of T-4-Hyp, with particular emphasis on biosynthetic pathways and core metabolic engineering strategies. Key approaches discussed include: mining and engineering the key enzyme, i.e., proline-4-hydroxylase (P4H), to enhance catalytic efficiency; systematically rewiring host metabolic networks by strengthening precursor supply (i.e., L-proline, α-ketoglutarate and L-glutamate), blocking the competing and degradation pathways, and dynamically regulating central carbon metabolism; optimizing fermentation processes through cofactor engineering (e.g., NADPH supply), improving oxygen availability, and precise regulation of key cofactors such as Fe²⁺. Finally, this review outlines the future research directions, highlighting the need for continued exploration in novel enzyme discovery, advanced systems metabolic engineering, and integrated process intensification. The convergence of multi-disciplinary technologies is expected to further promote the industrialization of microbial T-4-Hyp production.