Mechanism for the Coupling of ATP Hydrolysis to the Conversion of 5-Formyltetrahydrofolate to 5,10-Methenyltetrahydrofolate

Huang, Teng; Schirch, Verne

doi:10.1074/jbc.270.38.22296

Cited by 20 publications

(19 citation statements)

References 27 publications

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“…Inasmuch as the extent of folate secretion by bacteria can vary with the growth rate, pH, and ionic strength (52), it may well be that the conditions in situ provoke even greater secretion per cell than we observed in vitro. Upon uptake by spirochetes, 5-formyl-THF is presumably converted to 5,10-methenyl-THF by the physiologically irreversible synthetase (EC 6.3.3.2), which is widely distributed in prokaryotes and eukaryotes (21) and which serves to usher 5-formyl-THF into THF-dependent C 1 interconversion pathways. Although the impetus for this study was to identify an in situ source of folate for symbiotic spirochetes, 5-formyl-THF secretion by ZFX strains may be beneficial to the termites as well.…”

Section: Discussionmentioning

confidence: 99%

Folate Cross-Feeding Supports Symbiotic Homoacetogenic Spirochetes

Graber

Breznak

2005

Appl Environ Microbiol

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Treponema primitia, an H 2 -consuming CO 2 -reducing homoacetogenic spirochete in termite hindguts, requires an exogenous source of folate for growth. Tetrahydrofolate (THF) acts as a C 1 carrier in CO 2 -reductive acetogenesis, a microbially mediated process important to the carbon and energy requirements of termites. To examine the hypothesis that other termite gut microbes probably supply some form of folate to T. primitia in situ, we used a bioassay to screen for and isolate folate-secreting bacteria from hindguts of Zootermopsis angusticollis, which is the host of T. primitia. Based on morphology, physiology, and 16S rRNA gene sequences, the major folate secretors were identified as strains of Lactococcus lactis and Serratia grimesii. During growth, these isolates secreted 5-formyl-THF at levels up to 146 ng/ml, and their cell-free culture fluids satisfied the folate requirement of T. primitia strains in vitro. Analysis of Z. angusticollis hindgut fluid revealed that 5-formyl-THF was the only detectable folate compound and occurred at an in situ concentration (1.3 g/ml) which was more than sufficient to support the growth of T. primitia. These results imply that cross-feeding of 5-formyl-THF by other community members is important for growth of symbiotic hindgut spirochetes and thus termite nutrition and survival.Spirochetes are a major component of the hindgut microbiota of termites, accounting for as many as 50% of all prokaryotic cells (37), but for many years their role in the gut remained obscure as none had ever been isolated and studied in vitro. Recently, the first pure cultures of the following termite gut spirochetes were obtained: Treponema primitia strains ZAS-1 and ZAS-2 and Treponema azotonutricium strain 29,30). These organisms are believed to be important for termite nutrition because of their ability to fix dinitrogen and produce acetate, a major carbon and energy source for termites. In fact, T. primitia is a homoacetogen and possesses the Wood-Ljungdahl (acetyl coenzyme A) pathway for CO 2 -reductive acetogenesis (14). This enables T. primitia to conserve energy by homoacetogenesis from H 2 plus CO 2 , as well as from carbohydrates and methoxylated benzenoids, and to use CO 2 as a carbon source for growth. The Wood-Ljungdahl pathway is undoubtedly important in situ, because a significant portion of acetate formed in the hindgut arises from CO 2 reduction (5, 56). Given the abundance and species diversity of treponemes in termite hindguts (31), it is likely that many of the other not-yet-cultivated spirochetes are also CO 2 -reducing acetogens. This notion is supported by the large number of formyltetrahydrofolate synthetase (a key enzyme of the WoodLjungdahl pathway) genes similar to those of T. primitia detected in hindguts of Zootermopsis angusticollis, the termite that is the host of T. primitia (45).During recent studies of the physiology and nutrition of T. primitia, cells were found to require folate for growth (14). Requirements for exogenous folate compounds are not uncommon among...

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Section: Discussionmentioning

confidence: 99%

Folate Cross-Feeding Supports Symbiotic Homoacetogenic Spirochetes

Graber

Breznak

2005

Appl Environ Microbiol

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“…However, many deep-branching bacteria and archaea show no gene for this reaction, while multiple lines of evidence indicate that THF nonetheless functions as a carbonfixation cofactor in these organisms [22]. In almost all cases where an ATP-dependent N 10 -formyl-THF synthase is absent, an ATP-dependent N 5 -formyl-THF cycloligase [223,224] is found. This is another case where a broad evolutionary context allows an alternate interpretation.…”

Section: Refinement Of Folate-c 1 Chemistry Maps Onto Lineage Divergementioning

confidence: 99%

“…It is plausible that the N 5 -formyl-THF cycloligase allows a formate incorporation pathway that is an evolutionary intermediate between the commonly recognized pathway using THF and its evolutionary derivative using H 4 MPT (see figure 5). The ATP-dependent cycloligase produces N 5 ,N 10 -methenyl-THF from N 5 -formyl-THF, which may potentially form spontaneously due to the higher N 5 -pKa [224]. ATP hydrolysis is thus specifically linked to the N 10 -carbon bond, which is the primary donor for carbon groups from folates.…”

Section: Refinement Of Folate-c 1 Chemistry Maps Onto Lineage Divergementioning

confidence: 99%

“…This is another case where a broad evolutionary context allows an alternate interpretation. N 5 -formyl-THF cycloligase was originally discovered in mammalian systems, where its function has been highly uncertain and hypothesized to be the salvage mechanism as part of a futile cycle [223,224], before being found to be widespread across the tree of life [22]. If we deduce by reconstruction, however, that ancestral folate chemistry operated in the fully reductive direction, and that in H 4 MPT systems formate is attached at the N 5 position, while in THF systems formate is attached at the N 10 position, the widespread distribution of the cycloligase takes on a different possible meaning.…”

Section: Refinement Of Folate-c 1 Chemistry Maps Onto Lineage Divergementioning

confidence: 99%

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The compositional and evolutionary logic of metabolism

2012

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Metabolism is built on a foundation of organic chemistry, and employs structures and interactions at many scales. Despite these sources of complexity, metabolism also displays striking and robust regularities in the forms of modularity and hierarchy, which may be described compactly in terms of relatively few principles of composition. These regularities render metabolic architecture comprehensible as a system, and also suggests the order in which layers of that system came into existence. In addition metabolism also serves as a foundational layer in other hierarchies, up to at least the levels of cellular integration including bioenergetics and molecular replication, and trophic ecology. The recapitulation of patterns first seen in metabolism, in these higher levels, motivates us to interpret metabolism as a source of causation or constraint on many forms of organization in the biosphere. Many of the forms of modularity and hierarchy exhibited by metabolism are readily interpreted as stages in the emergence of catalytic control by living systems over organic chemistry, sometimes recapitulating or incorporating geochemical mechanisms. We identify as modules, either subsets of chemicals and reactions, or subsets of functions, that are re-used in many contexts with a conserved internal structure. At the small molecule substrate level, module boundaries are often associated with the most complex reaction mechanisms, catalyzed by highly conserved enzymes. Cofactors form a biosynthetically and functionally distinctive control layer over the small-molecule substrate. The most complex members among the cofactors are often associated with the reactions at module boundaries in the substrate networks, while simpler cofactors participate in widely generalized reactions. The highly tuned chemical structures of cofactors (sometimes exploiting distinctive properties of the elements of the periodic table) thereby act as 'keys' that incorporate classes of organic reactions within biochemistry. Module boundaries provide the interfaces where change is concentrated, when we catalogue extant diversity of metabolic phenotypes. The same modules that organize the compositional diversity of metabolism are argued, with many explicit examples, to have governed long-term evolution. Early evolution of core metabolism, and especially of carbon-fixation, appears to have required very few innovations, and to have used few rules of composition of conserved modules, to produce adaptations to simple chemical or energetic differences of environment without diverse solutions and without historical contingency. We demonstrate these features of metabolism at each of several levels of hierarchy, beginning with the small-molecule metabolic substrate and network architecture, continuing with cofactors and key conserved reactions, and culminating in the aggregation of multiple diverse physical and biochemical processes in cells.Content from this work may be used under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 3.0 licence.

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“…The reaction is ATP dependent and is subject to feedback inhibition by the product (16)(17)(18). Previous studies using nuclear magnetic resonance spectroscopy and isotope exchange methods have shown that the reaction proceeds via the formation of two intermediates (19). A nucleophilic attack by the formyl group of 5-formyltetrahydrofolate on the g-phosphate of ATP results in the formation of the first N5-iminium phosphate intermediate.…”

Section: Introductionmentioning

confidence: 99%

Structural Basis for the Inhibition of Human 5,10-Methenyltetrahydrofolate Synthetase by N10-Substituted Folate Analogues

Song

et al. 2009

Cancer Research

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5,10-Methenyltetrahydrofolate synthetase (MTHFS) regulates the flow of carbon through the one-carbon metabolic network, which supplies essential components for the growth and proliferation of cells. Inhibition of MTHFS in human MCF-7 breast cancer cells has been shown to arrest the growth of cells. Absence of the three-dimensional structure of human MTHFS (hMTHFS) has hampered the rational design and optimization of drug candidates. Here, we report the structures of native hMTHFS, a binary complex of hMTHFS with ADP, hMTHFS bound with the N5-iminium phosphate reaction intermediate, and an enzyme-product complex of hMTHFS. The N5-iminium phosphate captured for the first time in our crystal structure unravels a unique strategy used by hMTHFS for recognition of the substrate and provides structural basis for the regulation of enzyme activity. Binding of N10-substituted folate analogues places Y152 in the middle of the channel connecting ATP binding site with the substrate binding pocket, precluding the positioning of ;-phosphate for a nucleophilic attack. Using the structures of hMTHFS as a guide, we have probed the role of residues surrounding the active site in catalysis by mutagenesis. The ensemble of hMTHFS structures and the mutagenesis data yield a coherent picture of the MTHFS active site, determinants of substrate specificity, and new insights into the mechanism of inhibition of hMTHFS.

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Mechanism for the Coupling of ATP Hydrolysis to the Conversion of 5-Formyltetrahydrofolate to 5,10-Methenyltetrahydrofolate

Cited by 20 publications

References 27 publications

Folate Cross-Feeding Supports Symbiotic Homoacetogenic Spirochetes

Folate Cross-Feeding Supports Symbiotic Homoacetogenic Spirochetes

The compositional and evolutionary logic of metabolism

Structural Basis for the Inhibition of Human 5,10-Methenyltetrahydrofolate Synthetase by N10-Substituted Folate Analogues

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