Abstract
NAD+ synthetase is an essential enzyme of de novo and recycling pathways of NAD+ biosynthesis in Mycobacterium tuberculosis but not in humans. This bifunctional enzyme couples the NAD+ synthetase and glutaminase activities through an ammonia tunnel but free ammonia is also a substrate. Here we show that the Homo sapiens NAD+ synthetase (hsNadE) lacks substrate specificity for glutamine over ammonia and displays a modest activation of the glutaminase domain compared to tbNadE. We report the crystal structures of hsNadE and NAD+ synthetase from M. tuberculosis (tbNadE) with synthetase intermediate analogues. Based on the observed exclusive arrangements of the domains and of the intra- or inter-subunit tunnels we propose a model for the inter-domain communication mechanism for the regulation of glutamine-dependent activity and NH3 transport. The structural and mechanistic comparison herein reported between hsNadE and tbNadE provides also a starting point for future efforts in the development of anti-TB drugs.
Publication types
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Comparative Study
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Research Support, N.I.H., Extramural
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Amide Synthases / chemistry
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Amide Synthases / genetics
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Amide Synthases / metabolism*
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Ammonia / metabolism*
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Bacterial Proteins / chemistry
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Bacterial Proteins / genetics
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Bacterial Proteins / metabolism*
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Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor / chemistry
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Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor / genetics
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Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor / metabolism*
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Catalytic Domain
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Glutaminase / chemistry
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Glutaminase / genetics
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Glutaminase / metabolism
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Glutamine / metabolism
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Humans
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Mycobacterium tuberculosis / chemistry
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Mycobacterium tuberculosis / enzymology*
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Mycobacterium tuberculosis / genetics
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Mycobacterium tuberculosis / metabolism
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NAD / metabolism
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Substrate Specificity
Substances
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Bacterial Proteins
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Glutamine
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NAD
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Ammonia
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Glutaminase
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Amide Synthases
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NAD+ synthase
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Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor
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NADSYN1 protein, human