Microbial cell factories have been constructed via synthetic biology to convert renewable feedstocks into high-value chemicals, constituting a central objective of green biomanufacturing. With the advent of the “third-generation biorefinery” concept, CO2 has been recognized as a representative sustainable C1 feedstock whose bioconversion can simultaneously address feedstock sustainability and contribute to greenhouse-gas mitigation (Liu et al., 2020). Current routes for CO2 valorization can be broadly categorized into chemical and biological conversion. Chemical approaches, including photocatalytic conversion (Ganji et al., 2023), electrocatalytic reduction (Yu et al., 2024), and thermocatalytic hydrogenation (Zhao et al., 2022), can be directly coupled to renewable electricity or solar energy and are well suited for efficient production of small-molecule C1 intermediates such as methanol and formate. Nevertheless, large-scale implementation is frequently constrained by catalyst stability, costly downstream separation, and competing side reactions; these limitations are particularly pronounced for multistep transformations and the construction of complex, highly functionalized products, where high selectivity and high yield are difficult to achieve concurrently. By contrast, biological conversion proceeds under mild conditions, enabling high chemoselectivity and a broad product scope, but is often limited by gas-liquid mass-transfer inefficiency, substrate or intermediate toxicity, cofactor and energy constraints (Table 1). To address these bottlenecks, strategies that couple chemical and biological conversion have increasingly been pursued, in which CO2 is first converted through chemical routes into liquid intermediates more compatible with microbial metabolism, such as methanol and formate, and is subsequently assimilated by engineered cell factories for high-selectivity synthesis of value-added products.

Methanol is widely exploited by microorganisms as a carbon and energy source due to (1) its abundance, low cost, and ready accessibility; (2) its liquid state at ambient temperature, which facilitates storage and transport; and (3) its high energy density and high degree of reduction (6.0 for methanol versus 4.0 for glucose). These attributes position methanol as a key feedstock for microbial production of diverse chemicals. As a mature commodity chemical, methanol production has undergone decades of technological refinement and process optimization, yielding substantial advances in feedstock selection, production modes, energy efficiency, and process intensification (Fasihi and Breyer, 2024). Natural gas, coal, and biomass are employed as the principal feedstocks for methanol synthesis, with natural gas accounting for over 70% of global output. In recent years, green routes that use CO2 as the carbon source have been advanced, including photocatalytic conversion (Ganji et al., 2023; Wu et al., 2019b), electrocatalytic reduction (Li et al., 2023; Wu et al., 2019a; Yu et al., 2024), and thermocatalytic hydrogenation (Navarro-Jaén et al., 2021; Zhang et al., 2022; Zhao et al., 2022). By 2024, global methanol capacity had reached 186 million tons, with a market size of US$32.32 billion.

Despite these advances, methanol-driven biomanufacturing has remained constrained by the limited availability of methanol bio-converting cell factories. Two overarching strategies are pursued: (1) natural methylotrophs are deployed as chassis for native methanol bio-converting cell factories, which typically achieve superior carbon utilization but are limited by incompletely characterized genetics, constrained genome-editing toolkits, and narrow product spectra (Carrillo et al., 2019); and (2) model microorganisms, such as Escherichia coli, are engineered to create synthetic methanol bio-converting cell factories, which benefit from well-defined genetics, mature editing platforms, and extensive metabolic control, yet still exhibit low methanol assimilation efficiency, long doubling times, and modest product titers (Reiter et al., 2024).

In this review, the advances are synthesized across four dimensions to guide the development of efficient methanol bio-converting cell factories (Fig. 1): (1) the types and applications of methanol assimilation pathways; (2) energy-supply mechanisms operative in methanol metabolism and their constraints; (3) mechanisms underlying methanol toxicity and corresponding mitigation strategies; and (4) progress in microbial synthesis of chemicals from methanol together with prospects for application.