Publications

• Metabolic pathway rerouting in Paraburkholderia rhizoxinica evolved long-overlooked derivatives of coenzyme F420.

  Braga D., Last D., Hasan M., Guo H., Leichnitz D., Uzum Z., Richter I., Schalk F., Beemelmanns C., Hertweck C., Lackner G.

  Coenzyme F₄₂₀ is a specialized redox cofactor with a negative redox potential. It supports biochemical processes like methanogenesis, degradation of xenobiotics, and the biosynthesis of antibiotics. Although well-studied in methanogenic archaea and actinobacteria, not much is known abou ... >> More

  Coenzyme F₄₂₀ is a specialized redox cofactor with a negative redox potential. It supports biochemical processes like methanogenesis, degradation of xenobiotics, and the biosynthesis of antibiotics. Although well-studied in methanogenic archaea and actinobacteria, not much is known about F₄₂₀ in Gram-negative bacteria. Genome sequencing revealed F₄₂₀ biosynthetic genes in the Gram-negative, endofungal bacterium <i>Paraburkholderia rhizoxinica</i>, a symbiont of phytopathogenic fungi. Fluorescence microscopy, high-resolution LC-MS, and structure elucidation by NMR demonstrated that the encoded pathway is active and yields unexpected derivatives of F₄₂₀ (3PG-F₄₂₀). Further analyses of a biogas-producing microbial community showed that these derivatives are more widespread in nature. Genetic and biochemical studies of their biosynthesis established that a specificity switch in the guanylyltransferase CofC reprogrammed the pathway to start from 3-phospho-d-glycerate, suggesting a rerouting event during the evolution of F₄₂₀ biosynthesis. Furthermore, the cofactor activity of 3PG-F₄₂₀ was validated, thus opening up perspectives for its use in biocatalysis. The 3PG-F₄₂₀ biosynthetic gene cluster is fully functional in <i>Escherichia coli</i>, enabling convenient production of the cofactor by fermentation. << Less

  ACS Chem. Biol. 14:2088-2094(2019) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.

• Molecular insights into the biosynthesis of the F420 coenzyme.

  Forouhar F., Abashidze M., Xu H., Grochowski L.L., Seetharaman J., Hussain M., Kuzin A., Chen Y., Zhou W., Xiao R., Acton T.B., Montelione G.T., Galinier A., White R.H., Tong L.

  Coenzyme F(420), a hydride carrier, is found in Archaea and some bacteria and has crucial roles in methanogenesis, antibiotic biosynthesis, DNA repair, and activation of antitubercular compounds. CofD, 2-phospho-l-lactate transferase, catalyzes the last step in the biosynthesis of F(420)-0 (F(420) ... >> More

  Coenzyme F(420), a hydride carrier, is found in Archaea and some bacteria and has crucial roles in methanogenesis, antibiotic biosynthesis, DNA repair, and activation of antitubercular compounds. CofD, 2-phospho-l-lactate transferase, catalyzes the last step in the biosynthesis of F(420)-0 (F(420) without polyglutamate), by transferring the lactyl phosphate moiety of lactyl(2)diphospho-(5')guanosine to 7,8-didemethyl-8-hydroxy-5-deazariboflavin ribitol (Fo). CofD is highly conserved among F(420)-producing organisms, and weak sequence homologs are also found in non-F(420)-producing organisms. This superfamily does not share any recognizable sequence conservation with other proteins. Here we report the first crystal structures of CofD, the free enzyme and two ternary complexes, with Fo and P(i) or with Fo and GDP, from Methanosarcina mazei. The active site is located at the C-terminal end of a Rossmann fold core, and three large insertions make significant contributions to the active site and dimer formation. The observed binding modes of Fo and GDP can explain known biochemical properties of CofD and are also supported by our binding assays. The structures provide significant molecular insights into the biosynthesis of the F(420) coenzyme. Large structural differences in the active site region of the non-F(420)-producing CofD homologs suggest that they catalyze a different biochemical reaction. << Less

  J. Biol. Chem. 283:11832-11840(2008) [PubMed] [EuropePMC]

• A revised biosynthetic pathway for the cofactor F420 in prokaryotes.

  Bashiri G., Antoney J., Jirgis E.N.M., Shah M.V., Ney B., Copp J., Stuteley S.M., Sreebhavan S., Palmer B., Middleditch M., Tokuriki N., Greening C., Scott C., Baker E.N., Jackson C.J.

  Cofactor F₄₂₀ plays critical roles in primary and secondary metabolism in a range of bacteria and archaea as a low-potential hydride transfer agent. It mediates a variety of important redox transformations involved in bacterial persistence, antibiotic biosynthesis, pro-drug activation a ... >> More

  Cofactor F₄₂₀ plays critical roles in primary and secondary metabolism in a range of bacteria and archaea as a low-potential hydride transfer agent. It mediates a variety of important redox transformations involved in bacterial persistence, antibiotic biosynthesis, pro-drug activation and methanogenesis. However, the biosynthetic pathway for F₄₂₀ has not been fully elucidated: neither the enzyme that generates the putative intermediate 2-phospho-L-lactate, nor the function of the FMN-binding C-terminal domain of the γ-glutamyl ligase (FbiB) in bacteria are known. Here we present the structure of the guanylyltransferase FbiD and show that, along with its archaeal homolog CofC, it accepts phosphoenolpyruvate, rather than 2-phospho-L-lactate, as the substrate, leading to the formation of the previously uncharacterized intermediate dehydro-F₄₂₀-0. The C-terminal domain of FbiB then utilizes FMNH₂ to reduce dehydro-F₄₂₀-0, which produces mature F₄₂₀ species when combined with the γ-glutamyl ligase activity of the N-terminal domain. These new insights have allowed the heterologous production of F₄₂₀ from a recombinant F₄₂₀ biosynthetic pathway in Escherichia coli. << Less

  Nat. Commun. 10:1558-1558(2019) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.

• Characterization of the 2-phospho-L-lactate transferase enzyme involved in coenzyme F(420) biosynthesis in Methanococcus jannaschii.

  Graupner M., Xu H., White R.H.

  The protein product of the Methanococcus jannaschii MJ1256 gene has been expressed in Escherichia coli, purified to homogeneity, and shown to be involved in coenzyme F(420) biosynthesis. The protein catalyzes the transfer of the 2-phospholactate moiety from lactyl (2) diphospho-(5')guanosine (LPPG ... >> More

  The protein product of the Methanococcus jannaschii MJ1256 gene has been expressed in Escherichia coli, purified to homogeneity, and shown to be involved in coenzyme F(420) biosynthesis. The protein catalyzes the transfer of the 2-phospholactate moiety from lactyl (2) diphospho-(5')guanosine (LPPG) to 7,8-didemethyl-8-hydroxy-5-deazariboflavin (Fo) with the formation of the L-lactyl phosphodiester of 7,8-didemethyl-8-hydroxy-5-deazariboflavin (F(420)-0) and GMP. On the basis of the reaction catalyzed, the enzyme is named LPPG:Fo 2-phospho-L-lactate transferase. Since the reaction is the fourth step in the biosynthesis of coenzyme F(420), the enzyme has been designated as CofD, the product of the cofD gene. The transferase requires Mg(2+) for activity, and the catalysis does not appear to proceed via a covalent intermediate. To a lesser extent CofD also catalyzes a number of additional reactions that include the formation of Fo-P, when the enzyme is incubated with Fo and GDP, GTP, pyrophosphate, or tripolyphosphate, and the hydrolysis of F(420)-0 to Fo. All of these side reactions can be rationalized as occurring by a common mechanism. CofD has no recognized sequence similarity to any previously characterized enzyme. << Less

  Biochemistry 41:3754-3761(2002) [PubMed] [EuropePMC]