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Curr Protoc Nucleic Acid Chem. 2019 Apr 5:e83. doi: 10.1002/cpnc.83. [Epub ahead of print]

Chemoenzymatic Preparation of 4'-Thioribose NAD.

Author information

1
Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, California.
2
Department of Chemistry, Dornsife College of Letters, Arts and Sciences, University of Southern California, Los Angeles, California.
3
Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, California.
4
Research Center for Liver Diseases, University of Southern California, Los Angeles, California.

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.

KEYWORDS:

4′-thioribose; NAD+; NMNAT; NRK; chemoenzymatic synthesis; nicotinamide riboside

PMID:
30951610
DOI:
10.1002/cpnc.83

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