Relative Promiscuity Between Bacterial and Human Fad Synthetase: A Potential Source of Biomarkers

Abstract

FAD Synthetase (FADS) is a ubiquitous enzyme responsible for synthesizing flavin adenine dinucleotide (FAD) by the adenylation of flavin mononucleotide (FMN) using adenosine triphosphate (ATP). In bacteria, this enzyme is bifunctional, in that it catalyzes both the phosphorylation of riboflavin to produce flavin mononucleotide (FMN) and the adenylylation of FMN to produce flavin adenine dinucleotide (FAD). In contrast, the eukaryotic FADS enzyme is monofunctional, being capable of only the adenylation of FMN to form FAD. Studies on bacterial FADS have shown considerable promiscuity in the acceptance of altered FMNs as substrates but absolutely no uptake of different nucleotide triphosphates (e.g. CTP, TTP, etc.).Here we show that bacterial FADS (from C. ammoniagenes, CaFADS) will accept a modified ATP analog, specifically the fluorescent ATP analog aminopurine-2'-deoxyriboside-5'-triphosphate (2ApTP) for the production of dual fluorescent F-2Ap-D. Surprisingly, preliminary experiments indicate the monofunctional hFADS is not similarly promiscuous. These results implicate that this difference in substrate specificity may lead to unique biomarker applications. Understanding these enzymes may also lead to new approaches in nucleoside analog therapeutics.