AN 125I-LABELED N6-SUBSTITUTED AZIDO ANALOG OF NAD+ FOR THE PHOTOAFFINITY LABELING OF NAD+-LINKED ENZYMES |
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Authors: | Pei-Lin Chen Charles Mark Ensor Hsin-Hsiung Tai |
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Affiliation: | Division of Medicinal Chemistry &Pharmaceutics, College of Pharmacy, University of Kentucky, Lexington, KY 40536–0082, USA |
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Abstract: | Abstract 125I- N 6-(N-[6-N-{5-iodo-4-azidosalicyl}-aminohexyl]-aminocarbamoylmethyl)-nicotinamide adenine dinucleotide (125I- N 6-I-ASA-AH-NAD+) was synthesized by coupling N 6 -([6-aminohexyl]-carbamoylmethyl)-NAD+ with 4-azidosalicylic acid N-hydroxysuccinimide ester followed by radioiodination. The utility of 125I-N 6 -I-ASA-AH-NAD+ as an effective site-directed photoprobe was demonstrated by the photolabeling of both glutamate dehydro-genase and 15-hydroxyprostaglandin dehydrogenase. Both enzymes can be saturated with labeled probe with apparent dissociation constants comparable to those reported for NAD+. Photoincorporation of the probe into both enzymes was found to be protected specifically by NAD+. These results indicate that 125I- N 6-I-ASA-AH-NAD+ can be a specific photoprobe for NAD+-linked enzymes. |
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