Synthesis, Characterization, and Fluorescence Application of Indole-Based Isofunctional NAD+ Analogs
Danqi Li1, Bo Zhuang1, Ran-Ran Feng1
1Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
Researchers developed two fluorescent nicotinamide adenine dinucleotide (NAD+) analogs, 4CNI-NAD+ and 4CN7AI-NAD+, to overcome challenges in studying NAD+-dependent reactions. These analogs enable fluorescence-based monitoring of enzymatic processes involving NAD+ and NADH.
Area of Science:
- Biochemistry
- Molecular Biology
- Chemical Biology
Background:
- Nicotinamide adenine dinucleotide (NAD+) is crucial for cellular processes.
- Studying NAD+ reactions is difficult due to its nonfluorescent nature.
Purpose of the Study:
- To synthesize and characterize novel fluorescent NAD+ analogs.
- To enable fluorescence-based monitoring of NAD+-dependent enzymatic reactions.
Main Methods:
- Chemical synthesis of 4CNI-NAD+ and 4CN7AI-NAD+ analogs.
- Enzymatic assays using alcohol dehydrogenase (ADH) and NADase.
- Spectroscopic analysis (absorption and fluorescence).
- Molecular simulations and quantum chemical calculations.
Main Results:
- 4CNI-NAD+ and 4CN7AI-NAD+ mimic native NAD+ in biochemical properties.
- Enzyme reaction rates with 4CN7AI-NAD+ are comparable to NAD+.
- Reduced forms (NADH analogs) show decreased fluorescence.
- Nicotinamide cleavage significantly increases fluorescence.
- Photophysical properties are influenced by nicotinamide-cyanoindole interactions.
Conclusions:
- Developed fluorescent NAD+ analogs facilitate the study of NAD+-dependent enzymes.
- These analogs enable monitoring of NAD+/NADH hydrolysis via fluorescence changes.
- Expanded toolkit for investigating physiological NAD+-related processes.
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