Preugschat Frank, Tomberlin Ginger H, Porter David J T
Department of Biochemistry and Pharmacology, GlaxoSmithKline, 5 Moore Drive, 3.2114, Research Triangle Park, NC 27709, USA.
Arch Biochem Biophys. 2008 Nov 15;479(2):114-20. doi: 10.1016/j.abb.2008.09.005. Epub 2008 Sep 22.
ADP-ribosyl cyclase and NAD+ glycohydrolase (CD38, E.C.3.2.2.5) efficiently catalyze the exchange of the nicotinamidyl moiety of NAD+, nicotinamide adenine dinucleotide phosphate (NADP+) or nicotinamide mononucleotide (NMN+) with an alternative base. 4'-Pyridinyl drugs (amrinone, milrinone, dismerinone and pinacidil) were efficient alternative substrates (k(cat)/K(M)=0.9-10 microM(-1)s(-1)) in the exchange reaction with ADP-ribosyl cyclase. When CD38 was used as a catalyst the k(cat)/K(M) values for the exchange reaction were reduced two or more orders of magnitude (0.015-0.15 microM(-1)s(-1)). The products of this reaction were novel dinucleotides. The values of the equilibrium constants for dinucleotide formation were determined for several drugs. These enzymes also efficiently catalyze the formation of novel mononucleotides in an exchange reaction with NMN+, k(cat)/K(M)=0.05-0.4 microM(-1)s(-1). The k(cat)/K(M) values for the exchange reaction with NMN+ were generally similar (0.04-0.12 microM(-1)s(-1)) with CD38 and ADP-ribosyl cyclase as catalysts. Several novel heterocyclic alternative substrates were identified as 2-isoquinolines, 1,6-naphthyridines and tricyclic bases. The k(cat)/K(M) values for the exchange reaction with these substrates varied over five orders of magnitude and approached the limit of diffusion with 1,6-naphthyridines. The exchange reaction could be used to synthesize novel mononucleotides or to identify novel reversible inhibitors of CD38.
ADP-核糖基环化酶和NAD⁺糖水解酶（CD38，E.C.3.2.2.5）能高效催化NAD⁺、烟酰胺腺嘌呤二核苷酸磷酸（NADP⁺）或烟酰胺单核苷酸（NMN⁺）的烟酰胺基部分与另一种碱基的交换。4'-吡啶基药物（氨力农、米力农、双嘧力农和匹莫齐特）是与ADP-核糖基环化酶进行交换反应时的高效替代底物（k(cat)/K(M)=0.9 - 10 μM⁻¹s⁻¹）。当使用CD38作为催化剂时，交换反应的k(cat)/K(M)值降低了两个或更多数量级（0.015 - 0.15 μM⁻¹s⁻¹）。该反应的产物是新型二核苷酸。测定了几种药物形成二核苷酸的平衡常数。这些酶还能在与NMN⁺的交换反应中高效催化新型单核苷酸的形成，k(cat)/K(M)=0.05 - 0.4 μM⁻¹s⁻¹。以CD38和ADP-核糖基环化酶作为催化剂时，与NMN⁺的交换反应的k(cat)/K(M)值通常相似（0.04 - 0.12 μM⁻¹s⁻¹）。几种新型杂环替代底物被鉴定为2-异喹啉类、1,6-萘啶类和三环碱基。与这些底物进行交换反应的k(cat)/K(M)值在五个数量级范围内变化，对于1,6-萘啶类接近扩散极限。该交换反应可用于合成新型单核苷酸或鉴定CD38的新型可逆抑制剂。
