Structural insights into domain movement and cofactor specificity of glutamate dehydrogenase from Corynebacterium glutamicum

Son, Hyeoncheol Francis; Kim, Il-Kwon; Kim, Kyungjin

doi:10.1016/j.bbrc.2015.02.109

Cited by 26 publications

(30 citation statements)

References 16 publications

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“…A single subunit of CgGDH consisted of domain I (catalytic domain), Met1‐Val207 and Asn372‐Ile447, and domain II (nucleotide‐binding domain), Arg208‐Ala371, similar to other prokaryotic GDHs . The CgGDH·2‐IG·NADP + structure was essentially the same as the reported structure of CgGDH·2‐OG·NADP + complex with root‐mean‐square deviation of 0.21Å for 447 Cα atoms . Although all six chains in the asymmetric unit bound an NADP + molecule, four chains, A, B, C, and F, were in closed forms while the other two chains, D and E, were in open conformations.…”

Section: Resultssupporting

confidence: 58%

“…The crystal structure of the CgGDH complex binding 2‐OG and NADPH was reported by Son et al . . Their structure contained two hexamers in the asymmetric unit.…”

Section: Resultsmentioning

confidence: 99%

“…The CgGDHÁ2-IGÁNADP + structure was essentially the same as the reported structure of CgGDHÁ2-OGÁNADP + complex with root-mean-square deviation of 0. 21 A for 447 Ca atoms [16]. Although all six chains in the asymmetric unit bound an NADP + molecule, four chains, A, B, C, and F, were in closed forms while the other two chains, D and E, were in open conformations.…”

Section: Overall Structure Of Cggdhmentioning

confidence: 99%

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Crystal structure of the 2‐iminoglutarate‐bound complex of glutamate dehydrogenase from Corynebacterium glutamicum

et al. 2017

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Section: Resultssupporting

confidence: 58%

“…The crystal structure of the CgGDH complex binding 2‐OG and NADPH was reported by Son et al . . Their structure contained two hexamers in the asymmetric unit.…”

Section: Resultsmentioning

confidence: 99%

Section: Overall Structure Of Cggdhmentioning

confidence: 99%

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Crystal structure of the 2‐iminoglutarate‐bound complex of glutamate dehydrogenase from Corynebacterium glutamicum

et al. 2017

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“…Numerous crystal structures of GluDHs have been reported, including the crystal structure of the Corynebacterium glutamicum-derived GluDH (CgGluDH, PDB ID: 5IJZ) in complex with its NADP + cofactor and a-KG substrate. [12] Because CgGluDH shares a 71.4% sequence identity with PpGluDH, we used this crystal structure (the closed conformation) for homology modeling of PpGluDH. NADP + and a-KG were retained in the homology model ( Figure 1A).…”

Section: Rational Design Of Ppgludh Mutagenesismentioning

confidence: 99%

“…These four residues have been reported to play important roles in substrate stabilization through the formation of hydrogen bonds with the g-carboxyl O atoms of the glutamate/a-KG moiety. [12,14] We speculated that they may also be involved in stabilizing PPO during the catalytic process. In addition, these residues also form hydrogen bonds among themselves, such as the S381-R208 and S381-T196 bonds.…”

Section: Construction and Activity Assays Of "Cave-tailored" Mutantsmentioning

confidence: 99%

Rational Molecular Engineering of Glutamate Dehydrogenases for Enhancing Asymmetric Reductive Amination of Bulky α‐Keto Acids

et al. 2018

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Glutamate dehydrogenases (GluDHs) are promising biocatalysts for the synthesis of chiral aamino acids by asymmetric reductive amination of a-keto acids. However, their strict substrate specificity limits their applications. To address this problem, we developed a molecular engineering method for GluDHs that enhances the asymmetric reductive amination of bulky a-keto acids. Based on rational design, a "cave" located in the active site pocket of PpGluDH (GluDH from Pseudomonas putida), which plays an essential role in substrate recognition, was tailored to facilitate the accepting of bulky substrates. Two mutants (A167G and V378A) were discovered to have significantly enhanced catalytic activity toward 2-oxo-4-[(hydroxy)(methyl)phosphinyl]butyric acid (PPO) and several other bulky substrates. This molecular engineering method was then applied to ten other GluDHs from different sources and with different properties. All engineered GluDHs acquired substantial improvements in PPO-oriented catalytic activity. The most efficient mutant of NADP + (nicotinamide adenine dinucleotide phosphate)-specific GluDHs showed up to 1820-fold increased activity and the specific activity reached 111.02 U/mg-protein. The NAD + (nicotinamide adenine dinucleotide)-specific GluDHs, which have no detectable wild type activity toward PPO, acquired a considerable level of activity (1.90-29.48 U/mg-protein). In batch production of L-phosphinothricin, these "cave-tailored" GluDHs exhibited markedly improved catalytic efficiencies compared with their wild types and ee values of > 99%. The space-time yields (STY) varied from 818.16 to 1482.96 g · L À1 · d À1 , suggesting potential practical applications of these mutants.

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Molecular insights into the inhibition of glutamate dehydrogenase by the dicarboxylic acid metabolites

et al. 2021

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Glutamate dehydrogenase (GDH) is a salient metabolic enzyme which catalyzes the NAD + -or NADP + -dependent reversible conversion of α-ketoglutarate (AKG) to L-glutamate; and thereby connects the carbon and nitrogen metabolism cycles in all living organisms. The function of GDH is extensively regulated by both metabolites (citrate, succinate, etc.) and non-metabolites (ATP, NADH, etc.) but sufficient molecular evidences are lacking to rationalize the inhibitory effects by the metabolites. We have expressed and purified NADP + -dependent Aspergillus terreus GDH (AtGDH) in recombinant form. Succinate, malonate, maleate, fumarate, and tartrate independently inhibit the activity of AtGDH to different extents. The crystal structures of AtGDH complexed with the dicarboxylic acid metabolites and the coenzyme NADPH have been determined. Although AtGDH structures are not complexed with substrate; surprisingly, they acquire super closed conformation like previously reported for substrate and coenzyme bound catalytically competent Aspergillus niger GDH (AnGDH). These dicarboxylic acid metabolites partially occupy the same binding pocket as substrate; but interact with varying polar interactions and the coenzyme NADPH binds to the Domain-II of AtGDH. The low inhibition potential of tartrate as compared to other dicarboxylic acid metabolites is due to its weaker interactions of carboxylate groups with AtGDH. Our results suggest that the length of carbon skeleton and positioning of the carboxylate groups of inhibitors between two conserved lysine residues at the GDH active site might be the determinants of their inhibitory potency. Molecular details on the dicarboxylic acid metabolites bound AtGDH active site architecture presented here would be applicable to GDHs in general.

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Structural insights into domain movement and cofactor specificity of glutamate dehydrogenase from Corynebacterium glutamicum

Cited by 26 publications

References 16 publications

Crystal structure of the 2‐iminoglutarate‐bound complex of glutamate dehydrogenase from Corynebacterium glutamicum

Crystal structure of the 2‐iminoglutarate‐bound complex of glutamate dehydrogenase from Corynebacterium glutamicum

Rational Molecular Engineering of Glutamate Dehydrogenases for Enhancing Asymmetric Reductive Amination of Bulky α‐Keto Acids

Molecular insights into the inhibition of glutamate dehydrogenase by the dicarboxylic acid metabolites

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