[39] Preparation, Characterization, and Coenzymic Properties of 5-Carba-5-deaza and 1-Carba-1-deaza Analogs of Riboflavin, FMN, and FAD

This chapter discusses the preparation, characterization, and some of the biological and coenzymic properties of the riboflavin analogs, 5-carba-5-deazariboflavin, and 1-carba-l-deazariboflavin at the riboflavin, flavin mononucleotide (FMN), and flavin adenine dinucleotide (FAD) levels. These two analogs have carbon substituted for nitrogen at either end of the redox-active ethylenediamine linkage of the isoalloxazine ring and have proven quite useful, in complementary ways, as probes of flavoenzyme mechanisms. Phosphorylation of 5-carba-5-deazariboflavin to yield 5-carbadeazaFMN can be accomplished either chemically or enzymically. The enzymic route, although limited in the amount of material that can conveniently be prepared, appears to be the preferred route in view of some unwanted phosphorylation at the 4-position of the ribityl side chain during chemical phosphorylation. Enzymic phosphorylation of 5-carba-deazariboflavin can be accomplished, either by using adenosine triphosphate (ATP):riboflavin-5′-phosphotransferase (EC 2.7.1.26) from rat liver purified through the fourth step in the purification scheme of McCormick or using the Brevibacterium ammoniagenes flavokinase-FAD synthetase complex. An alternate procedure for obtaining 5-carba-deazaFMN involves the cleavage of 5-carba-deazaFAD by phosphodiesterase.