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Synthesis of UDP-6-deoxy-6,6-difluoro-D-GlcNAc : a potential mechanism-based inhibitor of pseB Bassiri, Jackie Mehrnaz


Pseudaminic acid is a nine carbon sugar, similar to sialic acid, and has been found to play an important role in the biosynthesis of flagella in pathogenic bacteria such as C. jejuni and H. pylori. For these bacteria the flagella is crucial in motility and colonization of the host. Because this sugar is found only in bacteria, enzymes required for its biosynthesis serve as novel targets for antibiotic development. A UDP-GlcNAc dehydratase, pseB, catalyses the first step of pseudaminic acid biosynthesis in C. jejuni. This enzyme catalyzes the conversion of UDP-GlcNAc to UDP-6-deoxy-4-keto- HexNAc. The proposed mechanism of this conversion is composed of oxidation of the C4 hydroxyl resulting in the formation of a 4-keto group, then a dehydration across the bond between C5 and C6 producing a UDP-4-keto-5,6-ene-hexose, and finally reduction in which a hydride from the enzyme bound NADFt cofactor is transferred to C6, and C5 is reprotonated on the opposite face such that the stereocentre at C5 is inverted during this dehydration reaction. In 1998, the synthesis of CDP-6-deoxy-6,6-difluoro-D-glucose, 9, was published by Chang et al. This compound was the first mechanism-based inactivator known for a dehydratase enzyme, more specifically, CDP-D-glucose 4,6-dehydratase, isolated from Yersinia pseudotuberculosis. Based on this work, a 10-step synthesis of UDP-6-deoxy- 6,6-difluoro-D-GlcNAc 11 was completed which generated a potential mechanism-based inhibitor for pseB. [image in original] The preliminary enzyme kinetics performed did not support 11 as a mechanismbased inactivator of pseB. One possible explanation for this is based on the steric aspect of the inhibitor vs. the substrate. The two fluorine groups on the C-6 positions are possibly too bulky for the inhibitor to bind the enzyme active site and cause inactivation. Another reason might be that the H-bonding of the C-6 hydroxyl group of the substrate to the enzyme is crucial in substrate binding. When it is replaced with two fluorines the lack of this H-bonding does not allow binding of the inhibitor to the active site.

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