99421-19-9Relevant articles and documents
3-Deoxy-3-fluoropyridoxamine 5'-phosphate: Synthesis and chemical and biological properties of a coenzyme B6 analog
Pieper,Yang,Zhou,Liu
, p. 1809 - 1817 (2007/10/03)
The C-3 deoxygenation of CDP-6-deoxy-L-threo-D-glycero-4-hexulose is the key step in the biosynthesis of ascarylose which is a 3,6-dideoxy sugar found in the lipopolysaccharide of Yersinia pseudotuberculosis. This transformation, achieved by the catalysis of CDP-6-deoxy-L-threo-D-glycero-4-hexulose-3-dehydrase (E1) and CDP-6-deoxy-L-threo-D-glycero-4-hexulose-3-dehydrase reductase (E3), is initiated by a reversible dehydration followed by a stepwise electron transfer from NADH to reduce the resulting glucoseen-PMP adduct. An organic radical intermediate has been detected by EPR during E1-E3 catalysis, and its characteristics are consistent with a phenoxyl radical. Its formation has been hypothesized to involve a tautomerization of the glucoseen-PMP intermediate to a PMP-quinone methide species, which then serves as the electron acceptor during the reduction. In order to gain further experimental evidence supporting this proposed mechanism, the title compound (F-PMP) was synthesized and tested for its competence as a cofactor for the E1-E3 reaction. Upon incubation with F-PMP, no C-3 deoxysugar product could be detected in the mixture. This result initially appeared to support the prediction that the 3-F substituent would prevent the tautomerization and thus inhibit the subsequent reduction. However, further analysis showed that the catalysis was actually arrested at the dehydration step since no 18O was incorporated at C-3 of the recovered substrate when the reaction was conducted in [18O]H2O, and no tritium was released when [4-3H]F-PMP replaced PMP in the incubation. Interestingly, the pK(a) of the ring nitrogen (N-1) of F-PMP was found to be 2.91, a value drastically altered from the 8.74 of PMP itself. Since the catalytic role of B6 coenzymes is to act as an electron sink, storing the electrons that are later used for the cleavage and/or formation of covalent bonds, protonation at N-1 is clearly essential, as it allows the formation of a salt bridge/hydrogen bond with an active site aspartate residue and also enhances the electron-withdrawing capability of the pyridine ring. Because F-PMP is not expected to exist in the pyridinium form at neutral pH, its ability to promote 4'-H abstraction is hampered since the resulting anion cannot be delocalized by the cofactor. Although incubation of E1-E3 with this new coenzyme B6 analog fails to provide additional support for the mechanism of this unique deoxygenation process, the results reported herein, along with those deduced from studies of other 3-substituted PLP derivatives, illustrate the importance of having an intact 3-OH group of the coenzyme in PLP/PMP dependent catalysis.
