Independent catalysis of the short form HisG from <i>Lactococcus lactis</i>

Livingstone, Emma; Mittelstädt, Gerd; Given, Fiona; Parker, Emily J.

doi:10.1002/1873-3468.12277

Cited by 14 publications

(51 citation statements)

References 29 publications

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“…The spectra of reactions catalysed by PaHisGS ( Figure 5A) and by PaATPPRT ( Figure 5B) are essentially identical, and both differ from the control reaction which lacked PaHisGS ( Figure 5C). The most conspicuous difference, besides the increase in the inorganic phosphate peak at those recently published for LlHisGS, 22 and indicate that the prevailing assumption that HisGS is catalytically inactive in the absence of HisZ 11, 12 should be revised.…”

Section: Pahisgs Catalysis In the Absence Of Pahisz Pahisgs Displaysmentioning

confidence: 85%

“…15 A similar observation of independent catalysis has been recently reported for LlHisGS. 22 In order to gather evidence in addition to the increase in UV absorbance at 290 nm 1,17 that PaHisGS is catalysing the same reaction as PaATPPRT, both reactions were compared by 31 P-NMR spectroscopy ( Figure 5). Spectra of both reactions, control, ATP, PRPP, and inorganic phosphate, along with assignment of chemical shifts, are shown in Figure S5.…”

Section: Pahisgs Catalysis In the Absence Of Pahisz Pahisgs Displaysmentioning

confidence: 99%

“…22 Likewise, while mesophilic and thermophilic ATPPRT's have been characterised, 11,12 no psychrophilic example has been reported.…”

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confidence: 99%

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Kinetics and Structure of a Cold-Adapted Hetero-Octameric ATP Phosphoribosyltransferase

Stroek

Talbot²,

Glok³

et al. 2017

Biochemistry

112

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ATP-phosphoribosyltransferase (ATPPRT) catalyses the first step in histidine biosynthesis, the condensation of ATP and 5-phospho--D-ribosyl-1-pyrophosphate to generate N 1 -(5-phospho--D-ribosyl)-ATP and inorganic pyrophosphate. The enzyme is allosterically inhibited by histidine. Two forms of ATPPRT, encoded by the hisG gene, exist in nature, depending on the species. The long form, HisGL, is a single polypeptide chain with catalytic and regulatory domains. The short form, HisGS, lacks a regulatory domain, and cannot bind histidine. HisGS instead is found in complex with a regulatory protein, HisZ, constituting the ATPPRT holoenzyme. HisZ triggers HisGS catalytic activity while rendering it sensitive to allosteric inhibition by histidine. Until recently, HisGS was thought to be catalytically inactive without HisZ. Here, recombinant HisGS and HisZ from the psychrophilic bacterium Psychrobacter arcticus were independently overexpressed and purified. The crystal structure of P. arcticus ATPPRT was solved at 2.34-Å resolution, revealing an equimolar HisGS-HisZ hetero-octamer. Steady-state kinetics indicate that both ATPPRT holoenzyme and HisGS are catalytically active. Surprisingly, HisZ confers only a modest 2-to 4-fold increase in kcat.Reaction profiles for both enzymes are indistinguishable by 31 P-NMR, indicating that the same reaction is catalysed. Temperature dependence of kcat shows deviation from Arrhenius behaviour at 308 K with the holoenzyme. Interestingly, such deviation is detected only at 313 K with HisGS. Thermal denaturation by CD spectroscopy resulted in Tm's of 312 K and 316 K for HisZ and HisGS, respectively, suggesting that HisZ renders the ATPPRT complex more thermolabile. This is the first characterisation of a psychrophilic ATPPRT. 4Adenosine 5ʹ-triphosphate phosphoribosyltransferase (ATPPRT) (EC 2.4.2.17) catalyses the reversible Mg 2+ -dependent reaction between adenosine 5ʹ-triphosphate (ATP) and(PR-ATP) and inorganic pyrophosphate (PPi) (Scheme 1), the first step in histidine biosynthesis. 1 The chemical equilibrium of the reaction strongly favours reactants, 2 and the enzyme is allosterically inhibited by histidine. 1 In addition to being a model for understanding allostery, 2-4 ATPPRT is of biotechnological interest as a tool for histidine production, provided that histidine feedback inhibition can be overcome. [5][6][7] Two forms of ATPPRT, encoded by the hisG gene, are found in nature. Fungi, plants, and most bacteria possess a long, homo-hexameric protein, HisGL, each subunit consisting of two domains that make up the catalytic core and a C-terminal regulatory domain that mediates feedback inhibition by histidine. 8 Some bacteria and archaea have a short version of the protein, HisGS, which lacks the C-terminal regulatory domain and is insensitive to histidine. In these organisms, a catalytically inactive regulatory protein, HisZ, the product of the hisZ gene, is present. 9 HisZ, which shares a common ancestry with histidyl-tRNA synthetase (HisRS), binds HisGS to form ...

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Section: Pahisgs Catalysis In the Absence Of Pahisz Pahisgs Displaysmentioning

confidence: 85%

Section: Pahisgs Catalysis In the Absence Of Pahisz Pahisgs Displaysmentioning

confidence: 99%

See 1 more Smart Citation

Kinetics and Structure of a Cold-Adapted Hetero-Octameric ATP Phosphoribosyltransferase

Stroek

Talbot²,

Glok³

et al. 2017

Biochemistry

112

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show abstract

“…The hisG gene encodes two forms of ATPPRT, a long-form of the protein (HisGL) found in plants, fungi, and most bacteria, 13,15 and a short-form (HisGS) present in archaea and some bacteria. [16][17] HisGL is a homo-hexamer with each subunit consisting of two N-terminal domains containing the active site and a C-terminal allosteric domain responsible for histidine inhibition.…”

Section: Introductionmentioning

confidence: 99%

Catalytic and Anticatalytic Snapshots of a Short-Form ATP Phosphoribosyltransferase

Alphey¹,

Fisher²,

Gould³

et al. 2018

ACS Catal.

101

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Allosteric modulation of catalysis is a common regulatory strategy of flux-controlling biosynthetic enzymes. The enzyme ATP phosphoribosyltransferase (ATPPRT) catalyses the first reaction in histidine biosynthesis, the magnesium-dependent condensation of ATP and 5phospho--D-ribosyl-1-pyrophosphate (PRPP) to generate N 1-(5-phospho--D-ribosyl)-ATP (PRATP) and pyrophosphate (PPi). ATPPRT is allosterically inhibited by the final product of the pathway, histidine. Hetero-octameric ATPPRT consists of four catalytic subunits (HisGS) and four regulatory subunits (HisZ) engaged in intricate catalytic regulation. HisZ enhances HisGS catalysis in the absence of histidine while mediating allosteric inhibition in its presence. Here we report HisGS structures for the apoenzyme and complexes with substrates (PRPP, PRPP-ATP, PRPP-ADP), product (PRATP), and inhibitor (AMP), along with ATPPRT holoenzyme structures in complexes with substrates (PRPP, PRPP-ATP, PRPP-ADP) and product (PRATP). These ten crystal structures provide an atomic view of the catalytic cycle and allosteric activation of Psychrobacter arcticus ATPPRT. In both ternary complexes with PRPP-ATP, the adenine ring is found in an anticatalytic orientation, rotated 180° from the catalytic rotamer. Arg32 interacts with phosphate groups of ATP and PRPP, bringing the substrates in proximity for catalysis. The negative charge repulsion is further attenuated by a magnesium ion sandwiched between the and -phosphate groups of both substrates. HisZ binding to form the hetero-octamer brings HisGS subunits closer together in a tighter dimer in the Michaelis complex, which poises Arg56 from the adjacent HisGS molecule for crosssubunit stabilisation of the PPi leaving group at the transition state. The more electrostatically pre-organised active site of the holoenzyme likely minimises the reorganisation energy required to accommodate the transition state. This provides a structural basis for allosteric activation in which chemistry is accelerated by facilitating leaving group departure.

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“…The upregulation of VdVA, VdATP6, and VdAC might also be related to ATP synthesis and the signal network [75][76][77]. When VdATP-PRT expression is strongly enhanced, glycometabolism may be stimulated and thus provide cellular energy [78][79][80]. Taken together, the accumulative evidence suggests that VdAK is a positive virulence regulator that is linked to energy metabolism.…”

Section: Discussionmentioning

confidence: 98%

Host-Induced Gene Silencing of an Adenylate Kinase Gene Involved in Fungal Energy Metabolism Improves Plant Resistance to Verticillium dahliae

et al. 2020

Biomolecules

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Verticillium wilt, caused by the ascomycete fungus Verticillium dahliae (Vd), is a devastating disease of numerous plant species. However, the pathogenicity/virulence-related genes in this fungus, which may be potential targets for improving plant resistance, remain poorly elucidated. For the study of these genes in Vd, we used a well-established host-induced gene silencing (HIGS) approach and identified 16 candidate genes, including a putative adenylate kinase gene (VdAK). Transiently VdAK-silenced plants developed milder wilt symptoms than control plants did. VdAK-knockout mutants were more sensitive to abiotic stresses and had reduced germination and virulence on host plants. Transgenic Nicotiana benthamiana and Arabidopsis thaliana plants that overexpressed VdAK dsRNAs had improved Vd resistance than the wild-type. RT-qPCR results showed that VdAK was also crucial for energy metabolism. Importantly, in an analysis of total small RNAs from Vd strains isolated from the transgenic plants, a small interfering RNA (siRNA) targeting VdAK was identified in transgenic N. benthamiana. Our results demonstrate that HIGS is a promising strategy for efficiently screening pathogenicity/virulence-related genes of Vd and that VdAK is a potential target to control this fungus.

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Independent catalysis of the short form HisG from Lactococcus lactis

Cited by 14 publications

References 29 publications

Kinetics and Structure of a Cold-Adapted Hetero-Octameric ATP Phosphoribosyltransferase

Kinetics and Structure of a Cold-Adapted Hetero-Octameric ATP Phosphoribosyltransferase

Catalytic and Anticatalytic Snapshots of a Short-Form ATP Phosphoribosyltransferase

Host-Induced Gene Silencing of an Adenylate Kinase Gene Involved in Fungal Energy Metabolism Improves Plant Resistance to Verticillium dahliae

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