Emoxipin

3-Hydroxypropionic acid (3-HP) is a versatile platform chemical with broad applications, serving as a precursor for the synthesis of value-added chemicals as well as the biodegradable polymers. However, current industrial production of 3-HP relies on chemical synthesis, which requires harmful raw materials and harsh reaction conditions. As a sustainable alternative, microbial biosynthesis of 3-HP has gained increasing attention. Yet, most reported pathways remain constrained by their dependence on vitamin B₁₂, a costly cofactor that limits scalability in industrial applications. Here, we report the development of a Corynebacterium glutamicum strain capable of high-level fermentative production of 3-HP from glucose via the introduction of a vitamin B₁₂-independent, β-alanine-derived pathway. Candidate genes for the conversion of β-alanine to 3-HP were first screened, and the optimized pathway was subsequently introduced into a previously developed β-alanine-overproducing BAL10 strain. By eliminating competing pathways to increase precursor availability, redirecting carbon flux through the pentose phosphate pathway to improve cofactor balance, strengthening the β-alanine biosynthetic pathway, and identifying a previously uncharacterized 3-HP transporter followed by fine-tuning its expression, the final engineered strain produced 126.3 g/L of 3-HP in high-inoculum fed-batch fermentation, with a yield of 0.36 g/g glucose and an overall productivity of 1.75 g/L/h. These results demonstrate the feasibility of a vitamin B₁₂-independent pathway for high-level 3-HP production, highlighting its potential for sustainable and scalable industrial application.