Thiamin Biosynthesis in Escherichia coli

Taylor, Sean V.; Kelleher, Neil L.; Kinsland, Cynthia; Chiu, Hsiu‐Ju; Costello, Colleen A.; Backstrom, Allyson D.; McLafferty, Fred W.; Begley, Tadhg P.

doi:10.1074/jbc.273.26.16555

Cited by 150 publications

(89 citation statements)

References 36 publications

(24 reference statements)

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“…To look for the putative intermediate (14), a sample of ThiF͞ThiS was rapidly purified to about 50% homogeneity from an overexpression strain (5). Analysis of this sample by using ESI͞FTMS indicated the presence of a new component of mass 34294.2-21 Da (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…A protein ThiS (12), which has been posttranslationally modified by conversion of its carboxyl-terminal glycine to a thiocarboxylate (ThiS-COSH), has been identified as an intermediate sulfur carrier (5,6), and the structure of this protein has been determined (7). Two enzymes involved in the formation of ThiS-COSH also have been identified and characterized: ThiF (5) catalyzes the formation of ThiS-COAMP, and IscS (8,9) catalyzes the sulfur transfer from cysteine to the acyl adenylate of ThiS in a pyridoxal 5Ј-phosphate-dependent reaction. Using both of these enzymes, ThiS-COSH formation has been successfully reconstituted in vitro (3,8).…”

Section: T Hiamin Pyrophosphate (3) Is An Essential Cofactor In All Lmentioning

confidence: 99%

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Biosynthesis of the thiazole moiety of thiamin in Escherichia coli : Identification of an acyldisulfide-linked protein–protein conjugate that is functionally analogous to the ubiquitin/E1 complex

Kinsland

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

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109

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A covalently linked protein-protein conjugate between ThiF and ThiS thiocarboxylate was found in a partially purified coexpressed ThiF͞ThiS protein mixture by using Fourier transform mass spectrometry. The Cys-184 of ThiF and the C terminus of ThiS thiocarboxylate were identified to be involved in the formation of this complex by using both mutagenesis and chemical modification methods. A complementation study of Escherichia coli thiF ؊ using thiF(C184S) suggests that this conjugate is an essential intermediate involved in the biosynthesis of the thiazole moiety of thiamin. This ThiF͞ThiS conjugate is the first characterized example of a unique acyldisulfide intermediate in a biosynthetic system. This protein conjugate is also an example of an ubiquitin-E1 like protein-protein conjugate in prokaryotes and supports a strong evolutionary link between thiamin biosynthesis and the ubiquitin conjugating system.

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Section: Resultsmentioning

confidence: 99%

Section: T Hiamin Pyrophosphate (3) Is An Essential Cofactor In All Lmentioning

confidence: 99%

Biosynthesis of the thiazole moiety of thiamin in Escherichia coli : Identification of an acyldisulfide-linked protein–protein conjugate that is functionally analogous to the ubiquitin/E1 complex

Kinsland

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

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109

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“…The closest analogies observed in nature are thiopeptide antibiotics like promoinducine (23) or thiostreptone (24) that are produced by Streptomyces species. Also, C-terminal amino acid sulfurylation is known to occur during the biosynthesis of thiamine (25) and molybdopterin (26).…”

Section: Discussionmentioning

confidence: 99%

The Biosynthesis of Methylated Amino Acids in the Active Site Region of Methyl-coenzyme M Reductase

Selmer¹,

Kahnt²,

Goubeaud³

et al. 2000

Journal of Biological Chemistry

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The global production of the greenhouse gas methane by methanogenic archaea reaches 1 billion tons per annum. The final reaction releasing methane is catalyzed by the enzyme methyl-coenzyme M reductase. The crystal structure of methyl-coenzyme M reductase from Methanobacterium thermoautotrophicum revealed the presence of five modified amino acids within the ␣-subunit and near the active site region. Four of these modifications were C-, N-, and S-methylations, two of which, 2-(S)-methylglutamine and 5-(S)-methylarginine, have never been encountered before. We have now confirmed these modifications by mass spectrometry of chymotryptic peptides. With methyl-coenzyme M reductase purified from cells grown in the presence of L-[methyl-D 3 ]methionine, it was shown that the methyl groups of the modified amino acids are derived from the methyl group of methionine rather than from methyl-coenzyme M, an intermediate in methane formation. The D 3 labeling pattern was found to be qualitatively and quantitatively the same as in the two methyl groups of the methanogenic coenzyme F 430 , which are known to be introduced via S-adenosylmethionine. From the results, it is concluded that the methyl groups of the modified amino acids in methyl-coenzyme M reductase are biosynthetically introduced by an S-adenosylmethioninedependent post-translational modification. A mechanism for the methylation of glutamine at C-2 and of arginine at C-5 is discussed.Methyl-coenzyme M reductase catalyzes the final reaction step in the formation of methane by methanogenic archaea. It is a 300-kDa protein composed of three different subunits in an ␣ 2 ␤ 2 ␥ 2 arrangement and contains per mol 2 mol of the nickel porphinoid coenzyme F 430 as prosthetic group (1, 2). The crystal structure of the enzyme from Methanobacterium thermoautotrophicum has recently been solved at 1.45-Å resolution (3). The electron density map suggested the presence of five modified amino acids in the ␣ subunits either at or very near the active site region: 1-N-methylhistidine ␣257 (3-methylhistidine according to IUPAC nomenclature), 5-(S)-methylarginine ␣271, 2-(S)-methylglutamine ␣400, S-methylcysteine ␣452, and thioglycine ␣445, forming a thiopeptide bond (Fig. 1). Thioglycine has been proposed to function as a one-electron relay in the catalytic mechanism (2). Methylation of histidine ␣257, which is involved in substrate binding, probably influences the substrate affinity of the enzyme (3). For the methylation of the three other amino acids, an accidental methylation via methylcoenzyme M-derived methyl radicals has been envisaged. We have now ruled out this possibility by showing that the methyl group of all four methylated amino acids is derived from the methyl group of methionine, most likely via S-adenosylmethionine (SAM).1 Our experiments are based on the previous finding that methionine is taken up from the medium by growing M. thermoautotrophicum and that the methionine taken up is used for protein synthesis and in SAM-dependent methylation reactions rather than in met...

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“…thiC and thiD are required for 4-amino-5-hydroxymethyl-2-methyl pyrimidine pyrophosphate synthesis (3, 4), whereas dxs (5, 6), thiF, thiS, thiG, thiH, and thiI are involved in 5-(2-hydroxyethyl)-4-methyl thiazole monophosphate synthesis (3,7,8). In addition, the product of the thiE gene couples 4-amino-5-hydroxymethyl-2-methyl pyrimidine pyrophosphate and 5-(2-hydroxyethyl)-4-methyl thiazole monophosphate to give thiamin monophosphate, which undergoes another phosphorylation catalyzed by the product of the thiL gene to form TPP, the biologically active form of vitamin B 1 (9, 10).…”

mentioning

confidence: 99%

A conserved RNA structure ( thi box) is involved in regulation of thiamin biosynthetic gene expression in bacteria

Miranda-Ríos

Navarro²,

Soberón³

2001

Proc. Natl. Acad. Sci. U.S.A.

227

153

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The thiCOGE genes of Rhizobium etli code for enzymes involved in thiamin biosynthesis. These genes are transcribed with a 211-base untranslated leader that contains the thi box, a 38-base sequence highly conserved in the 5 regions of thiamin biosynthetic and transport genes of Gram-positive and Gram-negative organisms. A deletion analysis of thiC-lacZ fusions revealed an unexpected relationship between the degree of repression shown by the deleted derivatives and the length of the thiC sequences present in the transcript. Three regions were found to be important for regulation: (i) the thi box sequence, which is absolutely necessary for high-level expression of thiC; (ii) the region immediately upstream to the translation start codon of thiC, which can be folded into a stem-loop structure that would mask the Shine-Dalgarno sequence; and (iii) the proximal part of the coding region of thiC, which was shown to contain a putative Rho-independent terminator. A comparative phylogenetic analysis revealed a possible folding of the thi box sequence into a hairpin structure composed of a hairpin loop, two helixes, and an interior loop. Our results show that thiamin regulation of gene expression involves a complex posttranscriptional mechanism and that the thi box RNA structure is indispensable for thiCOGE expression.T hiamin pyrophosphate (TPP), also known as cocarboxylase, is the cofactor of key enzymes of carbon metabolism such as pyruvate dehydrogenase, ␣-ketoglutarate dehydrogenase, transketolase, pyruvate decarboxylase, and others. TPP is made by the enzymatic coupling of two independently synthesized precursors, 4-amino-5-hydroxymethyl-2-methyl pyrimidine pyrophosphate and 5-(2-hydroxyethyl)-4-methyl thiazole monophosphate (1).In Escherichia coli and Salmonella enterica serovar Typhimurium, several genes are known to participate in TPP biosynthesis (2). thiC and thiD are required for 4-amino-5-hydroxymethyl-2-methyl pyrimidine pyrophosphate synthesis (3, 4), whereas dxs (5, 6), thiF, thiS, thiG, thiH, and thiI are involved in 5-(2-hydroxyethyl)-4-methyl thiazole monophosphate synthesis (3,7,8). In addition, the product of the thiE gene couples 4-amino-5-hydroxymethyl-2-methyl pyrimidine pyrophosphate and 5-(2-hydroxyethyl)-4-methyl thiazole monophosphate to give thiamin monophosphate, which undergoes another phosphorylation catalyzed by the product of the thiL gene to form TPP, the biologically active form of vitamin B 1 (9, 10). A salvage enzyme (thiamin kinase) encoded by thiK, which incorporates exogenous thiamin into TPP (11), also exists. In both organisms the thi genes are located throughout the chromosome and are arranged in three operons and four single gene loci (2). thiCEFSGH, thiMD (ThiM and ThiD are involved in salvage of 5-(2-hydroxyethyl)-4-methyl thiazole and 4-amino-5-hydroxymethyl-2-methyl pyrimidine from the culture medium, respectively; ThiD also catalyzes the phosphorylation of 4-amino-5-hydroxymethyl-2-methyl pyrimidine monophosphate and thiBPQ (which code for an ABC type transport system for t...

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Thiamin Biosynthesis in Escherichia coli

Cited by 150 publications

References 36 publications

Biosynthesis of the thiazole moiety of thiamin in Escherichia coli : Identification of an acyldisulfide-linked protein–protein conjugate that is functionally analogous to the ubiquitin/E1 complex

Biosynthesis of the thiazole moiety of thiamin in Escherichia coli : Identification of an acyldisulfide-linked protein–protein conjugate that is functionally analogous to the ubiquitin/E1 complex

The Biosynthesis of Methylated Amino Acids in the Active Site Region of Methyl-coenzyme M Reductase

A conserved RNA structure ( thi box) is involved in regulation of thiamin biosynthetic gene expression in bacteria

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