Corrinoid cofactor biosynthesis and insertion into reductive dehalogenases of anaerobic bacteria

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Corrinoid cofactors are formed in a protein-assisted multistep biosynthetic pathway. A unique corrinoid, norpseudo-B12, is a cofactor of the tetrachloroethene (PCE) reductive dehalogenase (PceA) of the gram-negative anaerobe Sulfurospirillum multivorans. PceA purified from the gram-positive anaerobe Desulfitobacterium hafniense strain PCE-S harbors a set of different corrinoid cofactors. The major goals of this project are to identify and characterize the diverse corrinoid cofactors present in reductive dehalogenases of anaerobic bacteria and to examine variations in the respective B12-biosynthetic pathways. UV/Vis-spectroscopy, mass spectrometric analyses, and nuclear magnetic resonance techniques will be used to characterize the isolated corrinoids. Genome mining, protein cross-linking, and interaction studies will help to identify proteins involved in the maturation of PceA. Mutagenesis of the respective genes will allow for a functional analysis of these proteins and their role in corrinoid biosynthesis and incorporation into the PceA precursor. Using subcellular localization techniques the membrane associated steps of corrinoid biosynthesis will be unravelled in this study. The project is meant to broaden the restricted knowledge on the biosynthesis and incorporation of corrinoid cofactors involved in the reductive dehalogenation by anaerobic bacteria.