Synthesis of N-acetylmannosamine-6-phosphate derivatives to investigate the mechanism of N-acetylmannosamine-6-phosphate 2-epimerase

• Published in: Carbohydrate research 2021-12, Vol.510, p.108445-108445, Article 108445
• Main Authors: Arif, Tanzeel, Currie, Michael J., Dobson, Renwick C.J., Newson, Harriet L., Poonthiyil, Vivek, Fairbanks, Antony J., North, Rachel A., Rendle, Phillip M.
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - chemistry; Bacterial Proteins - metabolism; Carbohydrate Conformation; Carbohydrate Epimerases - chemistry; Carbohydrate Epimerases - metabolism; Enzyme mechanism; Epimerase; Hexosamines - biosynthesis; Hexosamines - chemistry; N-Acetylglucosamine-6-phosphate; N-Acetylmannosamine-6-phosphate; NanE; Staphylococcus aureus - enzymology; Sugar Phosphates - biosynthesis; Sugar Phosphates - chemistry
• ISSN: 0008-6215; 1873-426X
• DOI: 10.1016/j.carres.2021.108445

The synthesis of analogues of natural enzyme substrates can be used to help deduce enzymatic mechanisms. N-Acetylmannosamine-6-phosphate 2-epimerase is an enzyme in the bacterial sialic acid catabolic pathway. To investigate whether the mechanism of this enzyme involves a re-protonation mechanism by the same neighbouring lysine that performed the deprotonation or a unique substrate-assisted proton displacement mechanism involving the substrate C5 hydroxyl, the syntheses of two analogues of the natural substrate, N-acetylmannosamine-6-phosphate, are described. In these novel analogues, the C5 hydroxyl has been replaced with a proton and a methyl ether respectively. As recently reported, Staphylococcus aureus N-acetylmannosamine-6-phosphate 2-epimerase was co-crystallized with these two compounds. The 5-deoxy variant bound to the enzyme active site in a different orientation to the natural substrate, while the 5-methoxy variant did not bind, adding to the evidence that this enzyme uses a substrate-assisted proton displacement mechanism. This mechanistic information may help in the design of potential antibacterial drug candidates. [Display omitted] •Synthesis of the 5-deoxy and 5-methoxy analogues of N-acetylmannosamine-6-phosphate.•Analogues provide evidence that N-acetylmannosamine-6-phosphate 2-epimerase uses substrate-assisted proton displacement.•Provides useful mechanistic information for the design of potential antibacterial drug candidates.