Mechanism of cyclizing NAD to cyclic ADP-ribose by ADP-ribosyl cyclase and CD38

Richard Graeff, Qun Liu, Irina A. Kriksunov, Masayo Kotaka, Norman Oppenheimer, Quan Hao, Hon Cheung Lee

Research output: Contribution to journalArticlepeer-review

53 Scopus citations

Abstract

Mammalian CD38 and its Aplysia homolog, ADP-ribosyl cyclase (cyclase), are two prominent enzymes that catalyze the synthesis and hydrolysis of cyclic ADP-ribose (cADPR), a Ca2+ messenger molecule responsible for regulating a wide range of cellular functions. Although both use NAD as a substrate, the cyclase produces cADPR, whereas CD38 produces mainly ADP-ribose (ADPR). To elucidate the catalytic differences and the mechanism of cyclizing NAD, the crystal structure of a stable complex of the cyclase with an NAD analog, ribosyl-2′F-2′-deoxynicotinamide adenine dinucleotide (ribo-2′-F-NAD), was determined. The results show that the analog was a substrate of the cyclase and that during the reaction, the nicotinamide group was released and a stable intermediate was formed. The terminal ribosyl unit at one end of the intermediate formed a close linkage with the catalytic residue (Glu-179), whereas the adenine ring at the other end stacked closely with Phe-174, suggesting that the latter residue is likely to be responsible for folding the linear substrate so that the two ends can be cyclized. Mutating Phe-174 indeed reduced cADPR production but enhanced ADPRproduction, converting the cyclase to be more CD38-like. Changing the equivalent residue in CD38, Thr-221 to Phe, correspondingly enhanced cADPR production, and the double mutation, Thr-221 to Phe and Glu-146 to Ala, effectively converted CD38 to a cyclase. This study provides the first detailed evidence of the cyclization process and demonstrates the feasibility of engineering the reactivity of the enzymes by mutation, setting the stage for the development of tools to manipulate cADPR metabolism in vivo.

Original languageEnglish (US)
Pages (from-to)27629-27636
Number of pages8
JournalJournal of Biological Chemistry
Volume284
Issue number40
DOIs
StatePublished - Oct 2 2009

Fingerprint

Dive into the research topics of 'Mechanism of cyclizing NAD to cyclic ADP-ribose by ADP-ribosyl cyclase and CD38'. Together they form a unique fingerprint.

Cite this