Scanning the active center of formolase to identify key residues for enhanced C1 to C3 bioconversion.

Background: Formolase (FLS) is a computationally designed enzyme that catalyzes the carboligation of two or three C1 formaldehyde molecules into C2 glycolaldehyde or C3 dihydroxyacetone (DHA). FLS lays the foundation for several artificial carbon fixation and valorization pathways, such as the artificial starch anabolic pathway. However, the application of FLS is limited by its low catalytic activity and product promiscuity.

Directed evolution of a neutrophilic and mesophilic methanol dehydrogenase based on high-throughput and accurate measurement of formaldehyde.

Methanol is a promising one-carbon feedstock for biomanufacturing, which can be sustainably produced from carbon dioxide and natural gas. However, the efficiency of methanol bioconversion is limited by the poor catalytic properties of nicotinamide adenine dinucleotide (NAD)-dependent methanol dehydrogenase (Mdh) that oxidizes methanol to formaldehyde. Herein, the neutrophilic and mesophilic NAD-dependent Mdh from DSM 2334 (Mdh) was subjected to directed evolution for enhancing the catalytic activity.

Engineering of the Translesion DNA Synthesis Pathway Enables Controllable C-to-G and C-to-A Base Editing in .

Expanding the base conversion type is expected to largely broaden the application of base editing, whereas it requires decipherment of the machinery controlling the editing outcome. Here, we discovered that the DNA polymerase V-mediated translesion DNA synthesis (TLS) pathway controlled the C-to-A editing by a glycosylase base editor (GBE) in . However, C-to-G conversion was surprisingly found to be the main product of the GBE in and subsequent gene inactivation identified the decisive TLS enzymes.