Mingli Zhao, Di Wu, Yuting Liao, Teng Chen, Wenli Zhang, Wanmeng Mu
L-cysteine is a naturally occurring sulfur-containing amino acid bearing a reactive thiol group. It plays vital roles in maintaining cellular redox homeostasis and synthesizing bioactive compounds, thus gaining widespread attention and broad applications in: food, pharmaceutical, cosmetic and feed industries. Currently, its industrial production mainly relies on keratin hydrolysis, which causes serious environmental and safety concerns. By contrast, biocatalytic and microbial fermentation have emerged as sustainable and safe alternatives. This review systematically summarizes current bioproduction strategies for L-cysteine, covering biocatalytic pathways and metabolic engineering of microbial cell factories. We further elaborate the metabolic pathways, regulatory networks, and transport systems of L-cysteine in three mainstream microbial chassis, including Escherichia coli, Corynebacterium glutamicum, and Pantoea ananatis. Core production bottlenecks and targeted metabolic engineering solutions are also comprehensively discussed. Finally, future prospects for sustainable industrial L-cysteine production are proposed.