Chemical and enzymic properties of riboflavin analogs

Abstract

The chemical and enzymatic properties of 26 analogues of riboflavin are presented. These analogues include both endo- and exocyclically substituted isoalloxazines with redox potentials from -370 to -128 mV. Physical and chemical data such as the electronic absorption spectra, pKas, and redox potentials of the analogues are presented and are discussed with respect to preferred tautomeric and resonance forms. Like riboflavin, most of the analogues are shown to be catalytic oxidants of dihydro-5-deazaflavins. Analogue binding to egg white binding apoprotein has been quantitated and serves to determine the origins of binding site specificity for this protein. Nearly all of the analogues that possess D-ribityl groups are found to be processed to the FAD level by the flavokinase/FAD synthetase system of Brevibacterium ammoniagenes. Most extensively studied are the reactivities of the analogues with the NAD(P)H:flavin oxidoreductase of Beneckea harveyi. Many of the analogues are substrates in this enzymatic redox reaction, and a linear free energy-rate relation (log Vmax vs. E0' of the analogue) is seen that parallels similar relationships in the nonenzymatic oxidation of dihydro-5-deazaflavins. This suggests a common mechanism for the reactions of such diverse flavins as riboflavin, 5-deazariboflavin, and 1-deazariboflavin.

Going through the Barrier

“…Substitution of the flavin prosthetic group in fulllength TbQSOX with 5-deaza-FAD (see "Experimental Procedures") yields an enzyme with undetectable activity (data not shown). This is to be expected because 5-deazaflavins are unreactive in their reduced forms toward oxygen (48). Although incubation of substituted TbQSOX with 5 mM DTT leads to insignificant reduction of the bound deazaflavin, the oxidized flavin envelope is blue shifted by ϳ5 nm (from 412 to 407 nm; Fig.…”

“…Because we wanted to determine the redox potential of the proximal disulfide in the context of an oxidized flavin, we explored the use of the highly reducing flavin analog 5-deaza-FAD (48,49). Substitution of the flavin prosthetic group in fulllength TbQSOX with 5-deaza-FAD (see "Experimental Procedures") yields an enzyme with undetectable activity (data not shown).…”

NAD(P)H‐flavin oxidoreductase from the bioluminescent bacterium, Vibrio fischeri ATCC 7744, is a flavoprotein

“…In this reaction, a free FMNH, binds to luciferase, which is provided by flavin reductase. The flavin reductase of luminous bacteria is quite unique in that it catalyzes a flavin redox reaction with flavins as substrate rather than as tightly bound flavins [8].…”

“…In this reaction, a free FMNH, binds to luciferase, which is provided by flavin reductase. The flavin reductase of luminous bacteria is quite unique in that it catalyzes a flavin redox reaction with flavins as substrate rather than as tightly bound flavins [8].Recently, the gene encoding NAD(P)H-flavin reductase was isolated from the bioluminescent bacterium, Vibrio jischeri ATCC 7744 (formerly Achromobacter jischeri or PhotobacteriumJischeri) [12]. Using the flavin reductase expressed in E. coli, it was possible to reconstitute the luminescence reaction in vitro with bacterial luciferase, the same as using native flavin reductase from J!…”

New flavins for old: artificial flavins as active site probes of flavoproteins

“…-370 mV (1,5-dideazaflavin) to -30 mV (8-phenylsulphonylflavin) (for useful tables of redox potentials see [27,[71][72][73]). Such an approach has been used to obtain convincing physical evidence that the long wavelength bands formed on adding phenols to Old Yellow Enzyme are due to charge transfer transitions involving the phenolate anion as donor and oxidized flavin as acceptor [72,74].…”