Fecha de publicación: -- Fuente: PubMed "industrial biotechnology" Bioresour Technol. 2026 Nov 9:136215. doi: 10.1016/j.biortech.2026.136215. Online ahead of print.ABSTRACTStreamlining metabolic pathways in microbes is a key strategy in metabolic engineering, often intended to channel evolution toward predictable outcomes. Here, Methylorubrum extorquens PA1 was rationally engineered by deleting redundant formate dehydrogenase operons (fdh234) to focus adaptive laboratory evolution on the remaining formate oxidation step under methanol stress. Genome sequencing identified a MetY-1 Y57D substitution in all five endpoint isolates from one evolving population, while the retained formate dehydrogenase operon carried no detected mutation. In vitro biochemical assays with purified MetY-1 showed that Y57D shifted the methoxine dose-response relative to the wild-type enzyme; at 5% (v/v) methanol the wild-type enzyme accumulated 0.386 ± 0.085 µM methoxine, whereas methoxine remained below the 0.025 µM detection limit for Y57D. A homocysteine-equivalent thiol signal, measured with Ellman reagent, remained detectable for both enzymes under the tested conditions. These results support a fine-tuning model for the product specificity of MetY-1 under methanol. The findings identify this specificity as a candidate point at which methanol toxicity may be mitigated; the contribution of the Y57D substitution to cellular growth was not tested here.PMID:43003316 | DOI:10.1016/j.biortech.2026.136215