Volume 77, Issue 1 e83
PROTOCOL

Chemoenzymatic Preparation of 4′-Thioribose NAD+

Xiao-Nan Zhang

Xiao-Nan Zhang

Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, California

Search for more papers by this author
Zhefu Dai

Zhefu Dai

Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, California

Search for more papers by this author
Qinqin Cheng

Qinqin Cheng

Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, California

Search for more papers by this author
Yong Zhang

Corresponding Author

Yong Zhang

Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, California

Department of Chemistry, Dornsife College of Letters, Arts and Sciences, University of Southern California, Los Angeles, California

Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, California

Research Center for Liver Diseases, University of Southern California, Los Angeles, California

Corresponding author: [email protected]Search for more papers by this author
First published: 05 April 2019
Citations: 1

Abstract

This chemoenzymatic procedure describes a strategy for the preparation of 4′-thioribose nicotinamide adenine dinucleotide (S-NAD+), including chemical synthesis of nicotinamide 4′-riboside (S-NR), recombinant expression and purification of two NAD+ biosynthesis enzymes nicotinamide riboside kinase (NRK) and nicotinamide mononucleotide adenylyltransferase (NMNAT), and enzymatic synthesis of S-NAD+. The first basic protocol describes the procedures for introduction of nicotinamide onto 4′-thioribose and subsequent deprotection to generate S-NR as the key intermediate for enzymatically synthesizing S-NAD+. In the second basic protocol, experimental methods are detailed for the production of recombinant human NRK1 and NMNAT1 to catalyze conversion of S-NR to S-NAD+. The third basic protocol presents the enzymatic approach for the generation of S-NAD+ from S-NR precursor. © 2019 by John Wiley & Sons, Inc.