Nickel-pincer nucleotide enzymes
镍钳核苷酸酶
基本信息
- 批准号:10656600
- 负责人:
- 金额:$ 34.51万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2019
- 资助国家:美国
- 起止时间:2019-07-01 至 2027-05-31
- 项目状态:未结题
- 来源:
- 关键词:AcidsAerococcusAnabolismBacteriaBacterial GenomeBindingBinding ProteinsBiochemicalBioinformaticsBiologicalC-terminalCarbonCarbon DioxideCatalysisCell WallCellsCoenzyme ACoenzymesComplementComplexCryoelectron MicroscopyCrystallographyDrug TargetingEnzymesGene ClusterGenesHealthHomologous GeneHumanHuman MicrobiomeHydrolysisHydroxy AcidsIn VitroInvestigationIronKnowledgeLactic acidLactobacillus plantarumLigand BindingLinkMetabolicMetabolismN-terminalNickelNucleotidesOrnithinePathogenicityPathway interactionsPropertyProtein Binding DomainProteinsPublic HealthRacemasesReactionResearchRoleSiteSpecificitySpectrophotometrySpectrum AnalysisStructureSulfidesSulfurSystemTestingThermotoga maritimaThioamidesVancomycin ResistanceVariantWorkZincadductanalogantimicrobialantimicrobial drugcarboxylatecarboxylationcofactorcysteine desulfurasedehydroalanineenantiomerenzyme structureepimerasehuman pathogenin vivointerestmicrobiomemicroorganismnicotinic acid adenine dinucleotidenovelpathogenpathogenic bacteriapersulfidesracemizationskills
项目摘要
Project Summary/Abstract
The nickel-pincer nucleotide (NPN) cofactor is a recently identified coenzyme discovered in lactate racemase
(LarA) but found more widely in 2-hydroxyacid racemases and epimerases. Synthesis of the cofactor requires
the sequential action of three accessory proteins: LarB carboxylates the pyridinium ring at C5 and hydrolyzes
the phosphoanhydride of nicotinic acid adenine dinucleotide (NaAD), LarE is an ATP-dependent enzyme that
converts the two pyridinium ring carboxylates into thiocarboxylates either by sacrificial loss of a protein cysteinyl
sulfur atom or by a mechanism involving a [4Fe-4S] cluster with a non-core sulfide, and LarC inserts nickel
(forming C-Ni and S-Ni bonds) in a CTP-dependent reaction. Gene homologs encoding these four proteins are
widely distributed in microorganisms associated with the human microbiome and among human pathogens. The
long-term objective of the effort described here is to significantly advance our understanding of how
microorganisms, including pathogenic species, make and utilize the NPN cofactor. Two specific aims will
achieve this objective: (1) the components of the NPN biosynthetic pathways will be structurally and
mechanistically characterized and (2) new roles of the NPN cofactor will be characterized beyond its function in
lactate racemase. Investigations of LarB will define the structure of an NaAD- and CO2-bound variant by
crystallography and confirm the formation of a covalent cysteinyl-pyridinium intermediate by stopped-flow UV-
vis spectrophotometry. In vivo analyses will test the cellular ability to reverse the sacrificial loss of cysteinyl sulfur
from one type of LarE. The [4Fe-4S] cluster-containing version of LarE will be structurally characterized by
crystallography. The hypothesis for a third, persulfide-containing form of LarE will be evaluated. The structure
and dynamics of intact LarC will be solved by cryo-electron microscopy. Alternatively, the N-terminal domain of
LarC will be defined by crystallography (complementing the known C-terminal structure) to better delineate the
enzyme’s single-turnover, CTP-dependent mechanism of nickel insertion. Additional LarA homologs will be
structurally and functionally defined. Examples that tightly bind larger substrates will be used to probe open
questions regarding the NPN-dependent enzyme reaction mechanism. Other examples are predicted to contain
both NPN and an iron-sulfur cluster whose function will be investigated. In addition, non-LarA-like proteins that
bind NPN will be identified and characterized. The findings obtained through these efforts will greatly increase
knowledge of the synthesis and utilization of nickel-pincer nucleotide cofactors in bacteria, including those
important to human health, with implications for identifying potential antimicrobial drug targets.
项目总结/文摘
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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ROBERT P HAUSINGER其他文献
ROBERT P HAUSINGER的其他文献
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{{ truncateString('ROBERT P HAUSINGER', 18)}}的其他基金
Characterization fo Fe(II)alpha-ketoglutarate-dependent hydroxylases
Fe(II)α-酮戊二酸依赖性羟化酶的表征
- 批准号:
7869489 - 财政年份:2009
- 资助金额:
$ 34.51万 - 项目类别:
Study of alpha-ketoglutarate-dependent dioxygenases
α-酮戊二酸依赖性双加氧酶的研究
- 批准号:
6472966 - 财政年份:2002
- 资助金额:
$ 34.51万 - 项目类别:
Study of alpha-ketoglutarate-dependent dioxygenases
α-酮戊二酸依赖性双加氧酶的研究
- 批准号:
6874849 - 财政年份:2002
- 资助金额:
$ 34.51万 - 项目类别:
Characterization of Fe(II)/alpha-ketoglutarate-dependent hydroxylases
Fe(II)/α-酮戊二酸依赖性羟化酶的表征
- 批准号:
8338809 - 财政年份:2002
- 资助金额:
$ 34.51万 - 项目类别:
Characterization of Fe(II)/alpha-ketoglutarate-dependent hydroxylases
Fe(II)/α-酮戊二酸依赖性羟化酶的表征
- 批准号:
8714000 - 财政年份:2002
- 资助金额:
$ 34.51万 - 项目类别:
Characterization fo Fe(II)alpha-ketoglutarate-dependent hydroxylases
Fe(II)α-酮戊二酸依赖性羟化酶的表征
- 批准号:
7596873 - 财政年份:2002
- 资助金额:
$ 34.51万 - 项目类别:
Characterization of Fe(II)/alpha-ketoglutarate-dependent hydroxylases
Fe(II)/α-酮戊二酸依赖性羟化酶的表征
- 批准号:
8538998 - 财政年份:2002
- 资助金额:
$ 34.51万 - 项目类别:














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