Abstract
Coenzyme F(420) plays a key role in the redox metabolisms of various archaea and bacteria, including Mycobacterium tuberculosis In M. tuberculosis, F(420)-dependent reactions have been linked to several virulence factors. F(420) carries multiple glutamate residues in the side chain, forming F(420)-n species (n, number of glutamate residues), and the length of this side chain impacts cellular physiology. M. tuberculosis strains with F(420) species carrying shorter side chains exhibit resistance to delamanid and pretomanid, two new tuberculosis (TB) drugs. Thus, the process of polyglutamylation of F(420) is of great interest. It has been known from genetic analysis that in mycobacteria an F(420)-0 γ-glutamyl ligase (FbiB) introduces up to seven glutamate residues into F(420) However, purified FbiB of M. tuberculosis (MtbFbiB) is either inefficient or incapable of incorporating more than two glutamates. We found that, in vitro, MtbFbiB synthesized side chains containing up to seven glutamate residues if F(420) was presented to the enzyme in a two-electron reduced state (F(420)H(2)). Our genetic analysis in Mycobacterium bovis BCG and Mycobacterium smegmatis and an analysis of literature data on M. tuberculosis revealed that in these mycobacteria the polyglutamylation process requires the assistance of F(420)-dependent glucose-6-phosphate dehydrogenase (Fgd) which reduces F(420) to F(420)H(2) We hypothesize that, starting with F(420)-0H(2), the amino-terminal domain of FbiB builds F(420)-2H(2), which is then transferred to the carboxy-terminal domain for further glutamylation; F(420)-2H(2) modifies the carboxy-terminal domain structurally to accommodate longer glutamyl chains. This system is analogous to folylpolyglutamate synthase, which introduces more than one glutamate residue into folate only after this vitamin is reduced to tetrahydrofolate.IMPORTANCE Coenzyme F(420)-dependent reactions of Mycobacterium tuberculosis, which causes tuberculosis, potentially contributes to the virulence of this bacterium. The coenzyme carries a glutamic acid-derived tail, the length of which influences the metabolism of M. tuberculosis Mutations that eliminate the production of F(420) with longer tails make M. tuberculosis resistant to two new tuberculosis drugs. This report describes that the synthesis of longer glutamyl tails of F(420) requires concerted actions of two enzymes, one of which reduces the coenzyme prior to the action of the other, which catalyzes polyglutamylation. This knowledge will help to develop more effective tuberculosis (TB) drugs. Remarkably, the introduction of multiple glutamate residues into the sidechain of folate (vitamin B(9)) requires similar concerted actions, where one enzyme reduces the vitamin to tetrahydrofolate and the other catalyzes polyglutamylation; folate is required for DNA and amino acid synthesis. Thus, the reported research has also revealed a key similarity between two important cellular systems.