Specific biotinylation of IMP dehydrogenase

Abstract

IMP dehydrogenase (IMPDH) catalyzes a critical step in guanine nucleotide biosynthesis. IMPDH also has biological roles that are distinct from its enzymatic function. We report a biotin-linked reagent that selectively labels IMPDH and is released by dithiothreitol. This reagent will be invaluable in elucidating the moonlighting functions of IMPDH. KeywordsIMPDH; enzyme inhibitors; nucleotide biosynthesis; 6-Cl-purine ribotide; biotin probe; mechanism-based probe Inosine 5'-monophosphate dehydrogenase (IMPDH) catalyzes the penultimate step of guanine nucleotide biosynthesis, the oxidation of IMP to XMP with the concomitant reduction of NAD +1 . IMPDH is up-regulated in rapidly dividing cells, and inhibition of IMPDH blocks proliferation and induces differentiation. IMPDH-targeted drugs are used in anticancer, antiviral, and immunosuppressive therapy, and are in development for antimicrobial applications. Some recent observations suggest additional functions for IMPDH beyond its well-established role in purine nucleotide biosynthesis. IMPDH associates with telomeric sequences, polyribosomes and actively transcribing promoters, localizes to lipid vesicles in response to insulin and aggregates in response to guanine nucleotide depletion [2][3][4][5][6] . Clearly many questions remain regarding the biological function of this housekeeping enzyme: is IMPDH enzymatically active in these different cellular contexts? what factors interact with IMPDH in each location? how do these interactions change in response to insulin and other signals?A mechanism-based probe would be an ideal tool to address these questions 7, 8. The IMPDH reaction is unusual among the enzymes involved in nucleotide metabolism in that it involves the nucleophillic attack of a conserved Cys residue, and several nucleotide analogs

Chiral Pool Synthesis of N-Cbz-cis-(3R,4R)-3-methylamino-4-methylpiperidine from L-Malic acid

“…[45][46][47] The direct methylation of amino piperidine compound 14 was not achievable for the reasons described before. 18 Reactions of amino piperidine 14 with p-anisaldehyde and formaldehyde under reductive conditions in DCM gave compound 16.…”