Extended Data Fig. 6: In vitro activity assay of METR-1.
From: Vitamin B12 produced by gut bacteria modulates cholinergic signalling
a, SDS–PAGE of purified METR-1 proteins. Purity of immunopurified GFP-tagged METR-1 protein was analysed with 4–10% polyacrylamide gels electrophoresis under denaturing conditions. Panels show silver staining (left) and western blot stained with a GFP antibody (right). Arrowhead indicates METR-1::GFP protein (molecular weight ~ 166 kDa). (n = 3 independent experiments with similar results) b, Methionine synthase activity of the purified METR-1::GFP protein was assayed in the presence of 5-methyltetrahydrofolate (5-meTHF) or betaine as a methyl donor ± SAM and DTT. Enzyme reaction was performed in 50 mM Tris-HCl buffer (pH 7.5) at 25 °C for 6 h (Methods) (mean ± s.e.m., one-way ANOVA with Dunnett’s multiple comparison). Source numerical data and source blot images are provided.
