Structures of the modified folates in the extremely thermophilic archaebacterium Thermococcus litoralis

White, Robert H.

doi:10.1128/jb.175.11.3661-3663.1993

Cited by 16 publications

(14 citation statements)

References 8 publications

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“…Hence, since UDP NAGs are the major, activated intermediates involved in the biosynthesis of both S-layers (22) and external structures, it may not be surprising that these compounds accumulate to such high intracellular concentrations in P. furiosus. In addition, both P. furiosus (44) and T. litoralis (43) have been shown to contain modified folates which have GlcNAc side chains attached. The reason for this is not known, but from a study (42) of methanopterin biosynthesis, it was concluded that GlcNAc residues are incorporated after folate synthesis.…”

Section: Vol 179 1997 Udp Amino Sugars In P Furiosus 1509mentioning

confidence: 99%

Characterization of UDP amino sugars as major phosphocompounds in the hyperthermophilic archaeon Pyrococcus furiosus

1997

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The archaeon Pyrococcus furiosus is a strictly anaerobic heterotroph that grows optimally at 100؇C by the fermentation of carbohydrates. It is known to contain high concentrations of novel intracellular solutes such as ␤-mannosylglycerate and di-myo-inositol 1,1-phosphate (DIP) (L. O. Martins and H. Santos, Appl. Environ. Microbiol. 61:3299-3303, 1995). Here, 31 P nuclear magnetic resonance (NMR) spectroscopy was used to show that this organism also accumulates another type of phospho compound, as revealed by a major multiplet signal in the pyrophosphate region. The compounds were purified from cell extracts of P. furiosus by anionexchange and gel filtration chromatographic procedures and were structurally analyzed by 1 H, 13 C, and 31 P NMR spectroscopy. They were identified as two uridylated amino sugars, UDP N-acetylglucosamine and UDP N-acetylgalactosamine. Unambiguous characterizations and complete assignments of 1 H and 13 C resonances from such sugars have not been previously reported. In vitro 31 P NMR spectroscopic analyses showed that, in contrast to DIP, which is maintained at a constant intracellular concentration (ϳ32 mM) throughout the growth phase of P. furiosus, the UDP amino sugars accumulated (to ϳ14 mM) only during the late log phase. The possible biochemical roles of these compounds in P. furiosus are discussed.Hyperthermophiles are a recently discovered group of microorganisms that have the unusual property of growing optimally at temperatures near and even above 100ЊC (7, 40). They have been isolated from geothermally heated environments, and most are of marine origin. Virtually all of them are classified under the domain Archaea rather than Bacteria (46). The domain Archaea includes some methanogenic and sulfate-reducing hyperthermophiles, but most of them are obligately anaerobic heterotrophs that reduce elemental sulfur (S 0 ) to H 2 S (2). The best studied of this group are members of the genus Pyrococcus (11). The type strain, Pyrococcus furiosus, is a marine organism that grows optimally at 100ЊC by the fermentation of peptides and carbohydrates. A variety of enzymes involved in the primary metabolic pathways of P. furiosus have been purified and characterized (1, 3).A key and, as yet, unresolved issue is how hyperthermophiles such as P. furiosus stabilize a wide range of biomolecules at extreme temperatures. This in turn has led to studies to determine if these organisms contain intracellular solutes that might have some general "thermoprotective" properties. This notion was first advanced by Scholz et al. (35), who found that Pyrococcus woesei maintains an intracellular potassium ion concentration of near 0.6 M and contains an equivalent concentration of the novel sugar di-myo-inositol-1,1Ј-phosphate (DIP) (Fig. 1). Moreover, under in vitro conditions, DIP was found to increase the thermal stability of an enzyme, glyceraldehyde-3-phosphate dehydrogenase, that has been purified from P. woesei (15,35). DIP is also present in the hyperthermophilic bacterium Thermotoga maritima (31) an...

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Section: Vol 179 1997 Udp Amino Sugars In P Furiosus 1509mentioning

confidence: 99%

Characterization of UDP amino sugars as major phosphocompounds in the hyperthermophilic archaeon Pyrococcus furiosus

1997

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“…In the case of coenzyme M, (S)-sulfolactate, formed from phosphoenolpyruvate (PEP) and bisulfite, is an intermediate in the biosynthesis (28-30). In the case of methanopterin (24, 25) and several related modified folates (33,34,36), (S)-hydroxyglutaric acid (23) is incorporated into the coenzyme during its biosynthesis (32, 35). For coenzyme F 420 (6) and its ␥-polyglutamate derivatives (7, 8, 18), (S)-hydroxypropionic acid (S-lactic acid) becomes a part of the final structure.…”

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Identification of an Archaeal 2-Hydroxy Acid Dehydrogenase Catalyzing Reactions Involved in Coenzyme Biosynthesis in Methanoarchaea

Graupner

White

2000

J Bacteriol

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Two putative malate dehydrogenase genes, MJ1425 and MJ0490, from Methanococcus jannaschii and one from Methanothermus fervidus were cloned and overexpressed in Escherichia coli, and their gene products were tested for the ability to catalyze pyridine nucleotide-dependent oxidation and reduction reactions of the following ␣-hydroxy-␣-keto acid pairs: (S)-sulfolactic acid and sulfopyruvic acid; (S)-␣-hydroxyglutaric acid and ␣-ketoglutaric acid; (S)-lactic acid and pyruvic acid; and 1-hydroxy-1,3,4,6-hexanetetracarboxylic acid and 1-oxo-1,3,4,6-hexanetetracarboxylic acid. Each of these reactions is involved in the formation of coenzyme M, methanopterin, coenzyme F 420 , and methanofuran, respectively. Both the MJ1425-encoded enzyme and the MJ0490-encoded enzyme were found to function to different degrees as malate dehydrogenases, reducing oxalacetate to (S)-malate using either NADH or NADPH as a reductant. Both enzymes were found to use either NADH or NADPH to reduce sulfopyruvate to (S)-sulfolactate, but the V max /K m value for the reduction of sulfopyruvate by NADH using the MJ1425-encoded enzyme was 20 times greater than any other combination of enzymes and pyridine nucleotides. Both the M. fervidus and the MJ1425-encoded enzyme catalyzed the NAD ؉ -dependent oxidation of (S)-sulfolactate to sulfopyruvate. The MJ1425-encoded enzyme also catalyzed the NADH-dependent reduction of ␣-ketoglutaric acid to (S)-hydroxyglutaric acid, a component of methanopterin. Neither of the enzymes reduced pyruvate to (S)-lactate, a component of coenzyme F 420 . Only the MJ1425-encoded enzyme was found to reduce 1-oxo-1,3,4,6-hexanetetracarboxylic acid, and this reduction occurred only to a small extent and produced an isomer of 1-hydroxy-1,3,4,6-hexanetetracarboxylic acid that is not involved in the biosynthesis of methanofuran c. We conclude that the MJ1425-encoded enzyme is likely to be involved in the biosynthesis of both coenzyme M and methanopterin.The biosynthesis of the methanogenic cofactors coenzyme M (2-mercaptoethanesulfonic acid), methanopterin, coenzyme F 420 , and methanofuran c (Fig. 1) requires the generation of an ␣-hydroxy acid that either becomes a component in the final structure or serves as an intermediate in the formation of the coenzyme. In the case of coenzyme M, (S)-sulfolactate, formed from phosphoenolpyruvate (PEP) and bisulfite, is an intermediate in the biosynthesis (28-30). In the case of methanopterin (24, 25) and several related modified folates (33,34,36), (S)-hydroxyglutaric acid (23) is incorporated into the coenzyme during its biosynthesis (32, 35). For coenzyme F 420 (6) and its ␥-polyglutamate derivatives (7, 8, 18), (S)-hydroxypropionic acid (S-lactic acid) becomes a part of the final structure. Finally, two (1R)-diastereomers of 1-hydroxy-1,3,4,6-hexanetetracarboxylic acid (HHTCA) serve as intermediates in the biosynthesis of the 1,3,4,6-hexanetetracarboxylic acid (HTCA) moiety of methanofuran (17; unpublished results), and another diastereomer of HHTCA [(1S)-HHTCA] is a component of metha...

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“…SHMT has been purified from both procaryotic and eucaryotic cells, and the cDNA sequences for more than 20 different forms are present in the SwissProt database. However, only limited information on SHMT from a member of the Archaea is available, in part because these organisms use modified folates as a coenzyme (13). These modified folates are not commercially available, making it difficult to assay for folate-requiring enzymes.…”

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“…The methanogens use methanopterin, a dimethylated pteridine (methyl groups at C-7 and C-9) connected to 1-(4-aminophenyl)-1-deoxy-D-ribitol (APDR), a common structural element found in all the currently known modified folates ( Fig. 1) (13,14). The hyperthermophile Sulfolobus solfataricus contains a modified folate which has structural features between those of the methanogenic modified folates and folate.…”

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Purification and properties of serine hydroxymethyltransferase from Sulfolobus solfataricus

et al. 1997

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Serine hydroxymethyltransferase (SHMT) catalyzes the reversible cleavage of serine to glycine with the transfer of the one-carbon group to tetrahydrofolate to form 5,10-methylenetetrahydrofolate. No SHMT has been purified from a nonmethanogenic Archaea strain, in part because this group of organisms uses modified folates as the one-carbon acceptor. These modified folates are not readily available for use in assays for SHMT activity. This report describes the purification and characterization of SHMT from the thermophilic organism Sulfolobus solfataricus. The exchange of the ␣-proton of glycine with solvent protons in the absence of the modified folate was used as the activity assay. The purified protein catalyzes the synthesis of serine from glycine and a synthetic derivative of a fragment of the natural modified folate found in S. solfataricus. Replacement of the modified folate with tetrahydrofolate did not support serine synthesis. In addition, this SHMT also catalyzed the cleavage of both allo-threonine and ␤-phenylserine in the absence of the modified folate. The cleavage of these two amino acids in the absence of tetrahydrofolate is a property of other characterized SHMTs. The enzyme contains covalently bound pyridoxal phosphate. Sequences of three peptides showed significant similarity with those of peptides of SHMTs from two methanogens.

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Structures of the modified folates in the extremely thermophilic archaebacterium Thermococcus litoralis

Cited by 16 publications

References 8 publications

Characterization of UDP amino sugars as major phosphocompounds in the hyperthermophilic archaeon Pyrococcus furiosus

Characterization of UDP amino sugars as major phosphocompounds in the hyperthermophilic archaeon Pyrococcus furiosus

Identification of an Archaeal 2-Hydroxy Acid Dehydrogenase Catalyzing Reactions Involved in Coenzyme Biosynthesis in Methanoarchaea

Purification and properties of serine hydroxymethyltransferase from Sulfolobus solfataricus

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