Molybdopterin is the essential component of the cofactors of all mononuclear molybdenum- and tungsten containing enzymes. Most procaryotic enzymes contain dinucleotide forms of molybdopterin with GMP, AMP, CMP or IMP. The most elaborate form of the cofactor is bis(molybdopterin guanine dinucleotide)-Mo, first identified in R. sphaeroides dimethyl sulfoxide reductase which contains no other cofactor. R. sphaeroides biotin sulfoxide reductase and E. coli trimethylamine-N- oxide reductase also contain bis(MGD)-Mo as the sole cofactor. These three-enzymes have been genetically cloned and have been successfully expressed in Escherichia coli. In collaboration with crystallographers elsewhere the X-ray structure of DMSO reductase has been determined, and we will attempt to obtain the crystal structures of the other two enzymes as well. Our ultimate goal is to examine in detail the structure- function interrelationship in these enzymes using a variety of spectroscopic and kinetic methods and availing site-directed mutants generated from a study of the crystal structure. The mechanism by which the complex bis(MGD)-Mo cofactor is assembled is also being investigated. We have cloned and have been able to overexpress many of the E. coli enzymes involved in the biosynthesis of molybdopterin and the variant forms of the cofactor. The crystal structures of two of the proteins, MoaC and MogA have been elucidated and will be used to explore the active sites and catalytic mechanisms. The reactions of the pathway will be studied in detail. These studies are of extreme importance, since molybdenum cofactor deficiency in humans causes severe neurological problems most often leading to neonatal death.
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| Nichols, Jason D; Xiang, Song; Schindelin, Hermann et al. (2007) Mutational analysis of Escherichia coli MoeA: two functional activities map to the active site cleft. Biochemistry 46:78-86 |
