Abstract text: 4-coumarate 3-hydroxylase/ascorbate peroxidase (C3H/APX) is a key gene in the lignin monomer biosynthesis pathway of monocotyledonous plants. Its activity relies on ascorbate or NADH as a cofactor; consequently, interfering with its expression in vivo often disrupts cofactor metabolism balance and leads to unpredictable phenotypic effects. Here, we report a novel, bacterial-derived C3H (nC3H) capable of directly converting 4-coumarate to caffeic acid without requiring ascorbate or NAD(P)H. Heterologous expression of nC3H in rice increased lignin content and altered lignin composition, resulting in enhanced biomass carbon content and improved disease resistance. Furthermore, we resolved the protein structure of nC3H at 2.5 Å resolution. Structure-guided engineering of residue Y88 produced a Y88F mutant with an approximately 10-fold increase in catalytic efficiency. Overall, this study provides new insights into lignin monomer biosynthesis and delivers an efficient enzymatic tool for targeted lignin modification.