title: Weak coupling of ATP hydrolysis to the chemical equilibrium of human nicotinamide phosphoribosyltransferase.
abstract: Human nicotinamide phosphoribosyltransferase (NAMPT, EC 2.4.2.12) catalyzes the reversible synthesis of nicotinamide mononucleotide (NMN) and inorganic pyrophosphate (PP i) from nicotinamide (NAM) and alpha- d-5-phosphoribosyl-1-pyrophosphate (PRPP). NAMPT, by capturing the energy provided by its facultative ATPase activity, allows the production of NMN at product:substrate ratios thermodynamically forbidden in the absence of ATP. With ATP hydrolysis coupled to NMN synthesis, the catalytic efficiency of the system is improved 1100-fold, substrate affinity dramatically increases ( K m (NAM) from 855 to 5 nM), and the K eq shifts -2.1 kcal/mol toward NMN formation. ADP-ATP isotopic exchange experiments support the formation of a high-energy phosphorylated intermediate (phospho-H247) as the mechanism for altered catalytic efficiency during ATP hydrolysis. NAMPT captures only a small portion of the energy generated by ATP hydrolysis to shift the dynamic chemical equilibrium. Although the weak energetic coupling of ATP hydrolysis appears to be a nonoptimized enzymatic function, closer analysis of this remarkable protein reveals an enzyme designed to capture NAM with high efficiency at the expense of ATP hydrolysis. NMN is a rate-limiting precursor for recycling to the essential regulatory cofactor, nicotinamide adenine dinucleotide (NAD (+)). NMN synthesis by NAMPT is powerfully inhibited by both NAD (+) ( K i = 0.14 muM) and NADH ( K i = 0.22 muM), an apparent regulatory feedback mechanism.
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461, USA.
authors.forename: Emmanuel S
authors.initials: ES
authors.lastname: Burgos
authors.forename: Vern L
authors.initials: VL
authors.lastname: Schramm
mesh_terms: D000178:Acrylamides / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000378:metabolism*; D016207:Cytokines / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D006801:Humans; D006868:Hydrolysis; D066298:In Vitro Techniques; D007700:Kinetics; D008954:Models, Biological; D009243:NAD / Q000378:metabolism / Q000494:pharmacology; D009537:Nicotinamide Mononucleotide / Q000378:metabolism; D054409:Nicotinamide Phosphoribosyltransferase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D010766:Phosphorylation; D010880:Piperidines / Q000494:pharmacology; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D013816:Thermodynamics
chemical_list: D000178:Acrylamides; D016207:Cytokines; D004791:Enzyme Inhibitors; C480543:N-(4-(1-benzoylpiperidin-4-yl)butyl)-3-(pyridin-3-yl)acrylamide; D010880:Piperidines; D011994:Recombinant Proteins; D009243:NAD; D009537:Nicotinamide Mononucleotide; D000255:Adenosine Triphosphate; D054409:Nicotinamide Phosphoribosyltransferase; C085455:nicotinamide phosphoribosyltransferase, human