Re-face stereospecificity of NADP dependent methylenetetrahydromethanopterin dehydrogenase from Methylobacterium extorquens AM1 as determined by NMR spectroscopy

MtdA catalyzes the dehydrogenation of N 5, N 10-methylenetetrahydromethanopterin (methylene-H 4MPT) with NADP + as electron acceptor. In the reaction two prochiral centers are involved, C14a of methylene-H 4MPT and C4 of NADP +, between which a hydride is transferred. The two diastereotopic protons at C14a of methylene-H 4MPT and at C4 of NADPH can be seen separately in 1H-NMR spectra. This fact was used to determine the stereospecificity of the enzyme. With (14a R)-[14a- 2H 1]-[14a- 13C]methylene-H 4MPT as the substrate, it was found that the pro-R hydrogen of methylene-H 4MPT is transferred by MtdA into the pro-R position of NADPH.