An Extended β7α7 Substrate-Binding Loop Is Essential for Efficient Catalysis by 3-Deoxy-<scp>d</scp>-<i>manno</i>-Octulosonate 8-Phosphate Synthase

Allison, Timothy M.; Hutton, Richard D.; Jiao, Wanting; Gloyne, B.J.; Nimmo, Evan B.; Jameson, Geoffrey B.; Parker, Emily J.

doi:10.1021/bi201231e

Cited by 5 publications

(8 citation statements)

References 33 publications

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“…Therefore, the modeled enzyme appeared to be a good starting point to study the interactions through docking and molecular dynamics simulations. Notably, the metal dependence of the KDO8P synthase has been extensively studied; recently, an evolutionary hypothesis showed that the catalytic activity of the metal-dependent KDO8P synthase is more compromised by the truncation of L7 than the metal-independent enzyme is [24]. The metal ion facilitates the correct coordination of A5P in the catalytic site; thus, metal independent enzymes are more reliant on the extended L7 loop for accurate A5P binding.…”

Section: Resultsmentioning

confidence: 99%

Computational Investigation of Bisphosphate Inhibitors of 3-Deoxy-d-manno-octulosonate 8-phosphate Synthase

et al. 2019

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The synthase, 3-deoxy-d-manno-octulosonate 8-phosphate (KDO8P), is a key enzyme for the lipopolysaccharide (LPS) biosynthesis of gram-negative bacteria and a potential target for developing new antimicrobial agents. In this study, computational molecular modeling methods were used to determine the complete structure of the KDO8P synthase from Neisseria meningitidis and to investigate the molecular mechanism of its inhibition by three bisphosphate inhibitors: BPH1, BPH2, and BPH3. Our results showed that BPH1 presented a protein–ligand complex with the highest affinity, which is in agreement with experimental data. Furthermore, molecular dynamics (MD) simulations showed that BPH1 is more active due to the many effective interactions, most of which are derived from its phosphoenolpyruvate moiety. Conversely, BPH2 exhibited few hydrogen interactions during the MD simulations with key residues located at the active sites of the KDO8P synthase. In addition, we hydroxylated BPH2 to create the hypothetical molecule named BPH3, to investigate the influence of the hydroxyl groups on the affinity of the bisphosphate inhibitors toward the KDO8P synthase. Overall, we discuss the main interactions between the KDO8P synthase and the bisphosphate inhibitors that are potential starting points for the design of new molecules with significant antibiotic activities.

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Section: Resultsmentioning

confidence: 99%

Computational Investigation of Bisphosphate Inhibitors of 3-Deoxy-d-manno-octulosonate 8-phosphate Synthase

et al. 2019

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“…2). This tetrameric assembly is a feature of type Iβ DAH7PS, which is shared by both regulated and unregulated DAH7PSs (19,21,23,37) and a related enzyme, 3-deoxy-D-manno-2-octulosonate 8-phosphate synthase (38). Intriguingly, quaternary structure is delicately balanced in this DAH7PS enzyme class.…”

Section: Discussionmentioning

confidence: 96%

Exploring modular allostery via interchangeable regulatory domains

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Jameson

et al. 2018

Proc. Natl. Acad. Sci. U.S.A.

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Most proteins comprise two or more domains from a limited suite of protein families. These domains are often rearranged in various combinations through gene fusion events to evolve new protein functions, including the acquisition of protein allostery through the incorporation of regulatory domains. The enzyme 3-deoxy-d--heptulosonate 7-phosphate synthase (DAH7PS) is the first enzyme of aromatic amino acid biosynthesis and displays a diverse range of allosteric mechanisms. DAH7PSs adopt a common architecture with a shared (β/α) catalytic domain which can be attached to an ACT-like or a chorismate mutase regulatory domain that operates via distinct mechanisms. These respective domains confer allosteric regulation by controlling DAH7PS function in response to ligand Tyr or prephenate. Starting with contemporary DAH7PS proteins, two protein chimeras were created, with interchanged regulatory domains. Both engineered proteins were catalytically active and delivered new functional allostery with switched ligand specificity and allosteric mechanisms delivered by their nonhomologous regulatory domains. This interchangeability of protein domains represents an efficient method not only to engineer allostery in multidomain proteins but to create a new bifunctional enzyme.

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“…The structures that are most similar to that of PaKDO8Ps were identified using the Structure Similarity option in PDBeFold (http:// www.ebi.ac.uk/msd-srv/ssm/). The highest structural similarity was found for the crystal structures of mutants of Neisseria meningitidis 3-deoxy-d-manno-octulosonate-8-phosphate synthase (NmKDO8Ps; Allison et al, 2011), which shares 68% sequence identity with PaKDO8Ps. Other known structures that share considerable sequence identity with PaKDO8Ps include the homologous enzymes from E. coli (PDB entry 1g7v; Asojo et al, 2001), with 69% sequence similarity, Burkholderia pseudomallei (PDB entry 3und; Seattle Structural Genomics Center for Infectious Disease, unpublished work), with 64% sequence identity, and Aquifex aeolicus (PDB entry 1fwt; Ackerman & Gatti, 2011), with 46% sequence identity.…”

Section: Comparison To Other Kdo8psmentioning

confidence: 99%

“…There are two classes of KDO8Ps: those that require a metal for catalysis and those that do not (Allison et al, 2011;Tao et al, 2010). Comparison of the PaKDO8Ps structure with homologous structures.…”

Section: Active Site Of Pakdo8psmentioning

confidence: 99%

Structure of 2-keto-3-deoxy-D-manno-octulosonate-8-phosphate synthase fromPseudomonas aeruginosa

et al. 2013

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Pseudomonas aeruginosa is a major cause of opportunistic infection and is resistant to most antibiotics. As part of efforts to generate much-needed new antibiotics, structural studies of enzymes that are critical for the virulence of P. aeruginosa but are absent in mammals have been initiated. 2-Keto-3-deoxy-D-manno-octulosonate-8-phosphate synthase (KDO8Ps), also known as 2-dehydro-3-deoxyphosphooctonate aldolase, is vital for the survival and virulence of P. aeruginosa. This enzyme catalyzes a key step in the synthesis of the lipopolysaccharide (LPS) of most Gram-negative bacteria: the condensation reaction between phosphoenolpyruvate (PEP) and arabinose 5-phosphate to produce 2-keto-3-deoxy-D-manno-octulosonate-8-phosphate (KDO8P). This step is vital for the proper synthesis and assembly of LPS and the survival of P. aeruginosa. Here, the recombinant expression, purification and crystal structure of KDO8Ps from P. aeruginosa are presented. Orthorhombic crystals were obtained by vapor diffusion in sitting drops in the presence of 1 mM phosphoenlpyruvate. The structure reveals the prototypical α/β TIM-barrel structure expected from this family of enzymes and contains a tetramer in the asymmetric unit.

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An Extended β7α7 Substrate-Binding Loop Is Essential for Efficient Catalysis by 3-Deoxy-d-manno-Octulosonate 8-Phosphate Synthase

Cited by 5 publications

References 33 publications

Computational Investigation of Bisphosphate Inhibitors of 3-Deoxy-d-manno-octulosonate 8-phosphate Synthase

Computational Investigation of Bisphosphate Inhibitors of 3-Deoxy-d-manno-octulosonate 8-phosphate Synthase

Exploring modular allostery via interchangeable regulatory domains

Structure of 2-keto-3-deoxy-D-manno-octulosonate-8-phosphate synthase fromPseudomonas aeruginosa

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