THE ROAD LESS TRAVELLED: UTILIZATION OF FORMATE IN TWO BIOCHEMICAL REACTIONS IN GRAM-NEGATIVE BACTERIA
University of Kansas
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Formate is an important intermediate in a number of metabolic reactions. Most of the formate pool in cells is generated from the degradation of primary metabolites (glucose, pyruvate, amino acids) and further degraded into CO2 and CH4. Remaining formate is recycled and utilized in one-carbon metabolism where a one-carbon formyl group is integrated into nucleotide metabolism, protein synthesis, and generation of secondary metabolites such as siderophores. Here I provide two examples of distinct and unique enzymes involved in formate generation and metabolism: PvdF from Pseudomonas aeruginosa and RibB from Vibrio Cholerae. PvdF is one of two enzymes involved in generation of the formyl-hydroxyornithine (fOHOrn) moiety responsible for iron chelation in pyoverdin, a siderophore in Pseudomonas aeruginosa. Biochemical and structural studies suggest that PvdF is a unique new class of transformylase enzyme. PvdF catalyzes the movement of the formyl group from an N10 formyl-THF analogue to the substrate following the bireactant random substrate binding model. Structurally, PvdF has a transformylase fold with secondary structural element insertion so far unprecedented in the literature. RibB is involved in the biosynthesis of riboflavin, vitamin B2. This is a magnesium dependent enzyme that catalyzes the conversion of the sugar ribulose 5-phosphate (Ru5P), a product of the pentose phosphate pathway, into 3,4-dihydroxy-2-butanone 4-phosphate (DHBP). The reaction catalyzed by RibB is an unusual deformylation reaction in which the fourth carbon of the five-carbon sugar is removed as formate. According to the literature, RibB catalyzes this reaction in the presence of di-metal Mg2+ centers following 1,2-methyl shift, called a skeletal rearrangement mechanism. Our evidence, both biochemical and structural, suggests that RibB requires only one Mg2+ for catalysis. Furthermore, NMR and X-ray crystallography data point toward the formation of a 2-phosphoglycolic acid intermediate during RibB catalyzed reaction. These data suggest that a fragmentation mechanism, not a skeletal rearrangement, is the preferred mechanism of RibB catalysis.
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