Unexpected Abundance of Coenzyme F420-Dependent Enzymes in Mycobacterium tuberculosis and Other Actinobacteria

ABSTRACT Regimens targeting Mycobacterium tuberculosis, the causative agent of tuberculosis (TB), require long courses of treatment and a combination of three or more drugs. An increase in drug-resistant strains of M. tuberculosis demonstrates the need for additional TB-specific drugs. A notable feature of M. tuberculosis is coenzyme F420, which is distributed sporadically and sparsely among prokaryotes. This distribution allows for comparative genomics-based investigations. Phylogenetic profiling (comparison of differential gene content) based on F420 biosynthesis nominated many actinobacterial proteins as candidate F420-dependent enzymes. Three such families dominated the results: the luciferase-like monooxygenase (LLM), pyridoxamine 5′-phosphate oxidase (PPOX), and deazaflavin-dependent nitroreductase (DDN) families. The DDN family was determined to be limited to F420-producing species. The LLM and PPOX families were observed in F420-producing species as well as species lacking F420 but were particularly numerous in many actinobacterial species, including M. tuberculosis. Partitioning the LLM and PPOX families based on an organism's ability to make F420 allowed the application of the SIMBAL (sites inferred by metabolic background assertion labeling) profiling method to identify F420-correlated subsequences. These regions were found to correspond to flavonoid cofactor binding sites. Significantly, these results showed that M. tuberculosis carries at least 28 separate F420-dependent enzymes, most of unknown function, and a paucity of flavin mononucleotide (FMN)-dependent proteins in these families. While prevalent in mycobacteria, markers of F420 biosynthesis appeared to be absent from the normal human gut flora. These findings suggest that M. tuberculosis relies heavily on coenzyme F420 for its redox reactions. This dependence and the cofactor's rarity may make F420-related proteins promising drug targets.

[1]  L. Daniels,et al.  Structures of coenzyme F420 in Mycobacterium species , 2001, Archives of Microbiology.

[2]  F. Bossa,et al.  Structure and mechanism of Escherichia coli pyridoxine 5'-phosphate oxidase. , 2003, Biochimica et biophysica acta.

[3]  L. Daniels,et al.  Mycobacterium smegmatis mc2 155 fbiC and MSMEG_2392 are involved in triphenylmethane dye decolorization and coenzyme F420 biosynthesis. , 2007, Microbiology.

[4]  C. Nathan,et al.  The Proteasome of Mycobacterium tuberculosis Is Required for Resistance to Nitric Oxide , 2003, Science.

[5]  T. Baldwin,et al.  Crystal structure of the bacterial luciferase/flavin complex provides insight into the function of the beta subunit. , 2009, Biochemistry.

[6]  Christian Cambillau,et al.  Crystal structure of the conserved hypothetical protein Rv1155 from Mycobacterium tuberculosis , 2005, FEBS letters.

[7]  I. Kumagai,et al.  Novel FMN-binding protein from Desulfovibrio vulgaris (Miyazaki F). Cloning and expression of its gene in Escherichia coli. , 1994, The Journal of biological chemistry.

[8]  A. Yasui,et al.  DNA photoreactivating enzyme from the cyanobacterium Anacystis nidulans. , 1990, The Journal of biological chemistry.

[9]  Robert H. White,et al.  Identification of the 7,8-didemethyl-8-hydroxy-5-deazariboflavin synthase required for coenzyme F420 biosynthesis , 2003, Archives of Microbiology.

[10]  L. Debussche,et al.  Purification of the two-enzyme system catalyzing the oxidation of the D-proline residue of pristinamycin IIB during the last step of pristinamycin IIA biosynthesis , 1995, Journal of bacteriology.

[11]  B. Mukhopadhyay,et al.  Conversion of NO2 to NO by reduced coenzyme F420 protects mycobacteria from nitrosative damage , 2009, Proceedings of the National Academy of Sciences.

[12]  F. Brockman,et al.  Cloning, sequencing, and analysis of a gene cluster from Chelatobacter heintzii ATCC 29600 encoding nitrilotriacetate monooxygenase and NADH:flavin mononucleotide oxidoreductase , 1997, Journal of bacteriology.

[13]  P. Wilairat,et al.  A novel two-protein component flavoprotein hydroxylase. , 2001, European journal of biochemistry.

[14]  Ross Upshur,et al.  XDR-TB in South Africa: No Time for Denial or Complacency , 2007, PLoS medicine.

[15]  J S Blanchard,et al.  Drugs that inhibit mycolic acid biosynthesis in Mycobacterium tuberculosis. , 2002, Current pharmaceutical biotechnology.

[16]  L. Daniels,et al.  Molecular Analysis of the Gene Encoding F420-Dependent Glucose-6-Phosphate Dehydrogenase fromMycobacterium smegmatis , 1998, Journal of bacteriology.

[17]  A. Halpern,et al.  The Sorcerer II Global Ocean Sampling Expedition: Northwest Atlantic through Eastern Tropical Pacific , 2007, PLoS biology.

[18]  L. Daniels,et al.  Structures of coenzyme F(420) in Mycobacterium species. , 2001, Archives of microbiology.

[19]  Michelle G. Giglio,et al.  TIGRFAMs and Genome Properties: tools for the assignment of molecular function and biological process in prokaryotic genomes , 2006, Nucleic Acids Res..

[20]  L. Daniels,et al.  Presence of F420-dependent glucose-6-phosphate dehydrogenase in Mycobacterium and Nocardia species, but absence from Streptomyces and Corynebacterium species and methanogenic Archaea. , 1997, FEMS microbiology letters.

[21]  S. Shima,et al.  Coenzyme binding in F420-dependent secondary alcohol dehydrogenase, a member of the bacterial luciferase family. , 2004, Structure.

[22]  Naomi Ward,et al.  Exopolysaccharide-associated protein sorting in environmental organisms: the PEP-CTERM/EpsH system. Application of a novel phylogenetic profiling heuristic , 2006, BMC Biology.

[23]  M. Winkler,et al.  Expression, purification, and characterization of recombinant Escherichia coli pyridoxine 5'-phosphate oxidase. , 1998, Protein expression and purification.

[24]  S. Shima,et al.  Crystal structure of methylenetetrahydromethanopterin reductase (Mer) in complex with coenzyme F420: Architecture of the F420/FMN binding site of enzymes within the nonprolyl cis‐peptide containing bacterial luciferase family , 2005, Protein science : a publication of the Protein Society.

[25]  L. Daniels,et al.  Mycobacterium smegmatis mc 2 155 fbiC and MSMEG _ 2392 are involved in triphenylmethane dye decolorization and coenzyme F 420 biosynthesis , 2007 .

[26]  M. James,et al.  Structural refinement of the non-fluorescent flavoprotein from Photobacterium leiognathi at 1.60 A resolution. , 1995, Journal of molecular biology.

[27]  M. Daffé,et al.  Molecular dissection of the biosynthetic relationship between phthiocerol and phthiodiolone dimycocerosates and their critical role in the virulence and permeability of Mycobacterium tuberculosis , 2007, The FEBS journal.

[28]  P. Bork,et al.  A human gut microbial gene catalogue established by metagenomic sequencing , 2010, Nature.

[29]  E. Eisenstein,et al.  Structure of the phenazine biosynthesis enzyme PhzG. , 2004, Acta crystallographica. Section D, Biological crystallography.

[30]  Pilho Kim,et al.  PA-824 Kills Nonreplicating Mycobacterium tuberculosis by Intracellular NO Release , 2008, Science.

[31]  E. Baker,et al.  Crystal Structures of F420-dependent Glucose-6-phosphate Dehydrogenase FGD1 Involved in the Activation of the Anti-tuberculosis Drug Candidate PA-824 Reveal the Basis of Coenzyme and Substrate Binding* , 2008, Journal of Biological Chemistry.

[32]  L. Quadri,et al.  Identification of Phthiodiolone Ketoreductase, an Enzyme Required for Production of Mycobacterial Diacyl Phthiocerol Virulence Factors , 2005, Journal of bacteriology.

[33]  Robert H. White,et al.  Identification and characterization of the 2-phospho-L-lactate guanylyltransferase involved in coenzyme F420 biosynthesis. , 2008, Biochemistry.

[34]  M. Kertesz,et al.  A Novel Reduced Flavin Mononucleotide-Dependent Methanesulfonate Sulfonatase Encoded by the Sulfur-Regulatedmsu Operon of Pseudomonas aeruginosa , 1999, Journal of bacteriology.

[35]  Rodrigo Lopez,et al.  Clustal W and Clustal X version 2.0 , 2007, Bioinform..

[36]  Robert H. White,et al.  Molecular Insights into the Biosynthesis of the F420 Coenzyme* , 2008, Journal of Biological Chemistry.

[37]  B. Roe,et al.  A core gut microbiome in obese and lean twins , 2008, Nature.

[38]  Clifton E. Barry,et al.  Tuberculosis — metabolism and respiration in the absence of growth , 2005, Nature Reviews Microbiology.

[39]  I. Sugawara,et al.  Overview of anti-tuberculosis (TB) drugs and their resistance mechanisms. , 2007, Mini reviews in medicinal chemistry.

[40]  Robert H. White,et al.  CofE catalyzes the addition of two glutamates to F420-0 in F420 coenzyme biosynthesis in Methanococcus jannaschii. , 2003, Biochemistry.

[41]  Robert C. Edgar,et al.  MUSCLE: a multiple sequence alignment method with reduced time and space complexity , 2004, BMC Bioinformatics.

[42]  J. Churchich.  Brain pyridoxine-5-phosphate oxidase. A dimeric enzyme containing one FMN site. , 1984, European journal of biochemistry.

[43]  Daniel H. Haft,et al.  Sites Inferred by Metabolic Background Assertion Labeling (SIMBAL): adapting the Partial Phylogenetic Profiling algorithm to scan sequences for signatures that predict protein function , 2010, BMC Bioinformatics.

[44]  Fan Yang,et al.  TIGRFAMs: a protein family resource for the functional identification of proteins , 2001, Nucleic Acids Res..

[45]  T. Bugg,et al.  Antimicrobial nucleoside antibiotics targeting cell wall assembly: recent advances in structure-function studies and nucleoside biosynthesis. , 2010, Natural product reports.