Emissive Synthetic Cofactors: Enzymatic Interconversions of ^tzA Analogues of ATP, NAD⁺, NADH, NADP⁺, and NADPH

Abstract

A series of enzymatic transformations, which generate visibly emissive isofunctional cofactors based on an isothiazolo[4,3-d]pyrimidine analogue of adenosine (^tz A), was developed. Nicotinamide adenylyl transferase condenses nicotinamide mononucleotide and ^tz ATP to yield N^tz AD⁺ , which can be enzymatically phosphorylated by NAD⁺ kinase and ATP or ^tz ATP to the corresponding N^tz ADP⁺ . The latter can be engaged in NADP-specific coupled enzymatic transformations involving conversion to N^tz ADPH by glucose-6-phosphate dehydrogenase and reoxidation to N^tz ADP⁺ by glutathione reductase. The N^tz ADP⁺ /N^tz ADPH cycle can be monitored in real time by fluorescence spectroscopy.