Glycerol kinase from Escherichia coli and an Ala65→Thr mutant: the crystal structures reveal conformational changes with implications for allosteric regulation

Feese, M.D.; Faber, H.R.; Bystrom, Cory; Pettigrew, Donald W.; Remington, S.J.

doi:10.1016/s0969-2126(98)00140-3

Cited by 56 publications

(65 citation statements)

References 55 publications

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“…6) about an axis through strands ␤11, ␤12, and ␤13 and two hinge loops that precede helix ␣23 (hinge loop-1) and strand ␤17 (hinge loop-2). This predicted movement is greater than in ecGK (22.3°) (43) and in Plasmodium falciparum glycerol kinase (27°) (46); superpositions show that the domains are wider apart in the open form of hXK. The modeled closure of hXK buries the Xu substrate and brings the ␥-phosphate of ATP in contact with the O5 hydroxyl of Xu.…”

Section: Resultsmentioning

confidence: 76%

“…Conformational Change-Enzymes of the sugar kinase/ hsp70/actin superfamily are known to undergo large scale domain movements that enable catalysis to occur; these movements have been well described, for example, for hexokinase (42) and glycerol kinase (43). The hXK structures determined here are all in an "open" form, in which the two domains are widely separated and the two substrates at least 10 Å apart.…”

Section: Resultsmentioning

confidence: 85%

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Structure and Function of Human Xylulokinase, an Enzyme with Important Roles in Carbohydrate Metabolism

Bunker¹,

Bulloch

Dickson

et al. 2013

Journal of Biological Chemistry

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Background: D-Xylulokinase (XK), the final enzyme in the glucuronate-xylulose pathway, produces a key regulator of lipogenesis, xylulose 5-phosphate. Results: The structure of human XK was determined, and its catalytic activity and inhibition were characterized. Conclusion: Human XK is selective for D-xylulose and is inhibited by 5-deoxy-5-fluoro-D-xylulose. Significance: Inhibition of XK could clarify its roles in sugar metabolism, lipogenesis, and metabolic disease.

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Section: Resultsmentioning

confidence: 76%

Section: Resultsmentioning

confidence: 85%

Structure and Function of Human Xylulokinase, an Enzyme with Important Roles in Carbohydrate Metabolism

Bunker¹,

Bulloch

Dickson

et al. 2013

Journal of Biological Chemistry

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“…Positions 427-429 are critical for the binding of IIA Glc and coupling to the active site. Crystal structures show that these amino acids do not form interactions with IIA Glc , and there is no change in conformation in this region after IIA Glc binding that is discernible at the 2.4-3.0-Å resolutions of the structures (7,10,11). No atoms of the amino acids at 427-429 are closer than 4.3 Å to IIA Glc , and no IIA Glc amino acid side chains form polar interactions with these amino acids.…”

Section: Resultsmentioning

confidence: 95%

“…Crystallographic studies show that the conformations of EcGK residues 474-478 undergo induced-fit transitions between coil, 3 10 helix, and ␣-helix upon IIA Glc binding (11). Alanine substitutions at positions 479-481, which are ␣-helical in the absence or presence of IIA Glc , result in large decreases in affinity that are consistent with stabilization of the ␣-helix, affecting positions that undergo the conformational changes (29).…”

Section: Resultsmentioning

confidence: 96%

“…Posttranslational regulation of the function of many of the family members is believed to involve modulation of this motion. Crystallographic studies of EcGK show evidence for conformational changes associated with active site closure and induced fit of amino acids that bind IIA Glc but do not show how these two conformational changes are coupled (10,11). The conformation of IIA Glc does not change after binding to EcGK.…”

mentioning

confidence: 91%

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Transplanting allosteric control of enzyme activity by protein–protein interactions: Coupling a regulatory site to the conserved catalytic core

Pawlyk

Pettigrew²

2002

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

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Glycerol kinase from Escherichia coli, but not Haemophilus influenzae, is inhibited allosterically by phosphotransferase system protein IIA Glc . The primary structures of these related kinases contain 501 amino acids, differing at 117. IIA Glc inhibition is transplanted from E. coli glycerol kinase into H. influenzae glycerol kinase by interconverting only 11 of the differences: 8 residues that interact with IIA Glc at the allosteric binding site and 3 residues in the conserved ATPase catalytic core that do not interact with IIA Glc but the solvent accessible surface of which decreases when it binds. The three core residues are crucial for coupling the allosteric site to the conserved catalytic core of the enzyme. The site of the coupling residues identifies a regulatory locus in the sugar kinase͞ heat shock protein 70͞actin superfamily and suggests relations between allosteric regulation and the active site closure that characterizes the family. The location of the coupling residues provides empirical validation of a computational model that predicts a coupling pathway between the IIA Glc A llosteric regulation of enzyme catalysis is a fundamental aspect of control in biological systems. For most classically studied allosteric enzymes (1), catalytic activity is modulated by small molecules such as fructose 1,6-bisphosphate (FBP), phosphoenolpyruvate, or nucleotides. The regulatory binding sites and the active sites are located at subunit-subunit interfaces of the homooligomeric proteins and are separated by Ͼ10 Å. Substrate binding shows homotropic interactions, typically positive cooperativity. Allosteric effectors change the affinity for substrates at the active sites but do not affect V max . Allosteric inhibition decreases apparent affinity for substrate, and activation increases apparent affinity. Changes in quaternary structure, involving small rigid-body rotations and translations of subunits, are seen in crystal structures of the enzymes with substrates or allosteric effectors. Regulation of catalysis by protein-protein interactions is less understood but seems to be the rule rather than the exception, and there is widespread interest in interactomes and roles of macromolecular interactions in living systems (2). A related area of interest is the molecular evolution of allosteric control by protein-protein interactions. Here we show that transplantation of allosteric regulation from one enzyme to another provides insights into how protein-protein interactions distant from an active site regulate catalysis and into mechanisms of molecular evolution.Glycerol kinase (EC 2.7.1.30; ATP:glycerol 3-phosphotransferase) from Escherichia coli (EcGK) is a regulatory enzyme (3). It is inhibited allosterically by IIA Glc , the 18-kDa glucose-specific phosphocarrier protein of the phosphoenolpyruvate:glycose phosphotransferase system (4). Binding of IIA Glc decreases V max and the substrate Michaelis constants, and no cooperativity is observed for inhibition or with respect to glycerol or MgATP (5).Thus, in contra...

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Rat Small Intestinal Mucosal Epithelial Cells Absorb Dietary 1,3‐Diacylglycerol Via Phosphatidic Acid Pathways

Zou

et al. 2018

Lipids

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Compared with triacylglycerol (TAG), dietary 1,3-diacylglycerol (1,3-DAG) is associated with reduced serum lipid and glucose levels. We investigated the metabolism of 1,3-DAG by assaying its intermediate metabolites during digestion and absorption in the rat small intestine. After gavage with TAG emulsion, TAG was digested mainly to 2-monoacylglycerol (2-MAG) and unesterified fatty acid (FFA) in the rat small intestinal lumen. 2-MAG was directly absorbed into the small intestinal epithelial cells and esterified to 1,2(2,3)-DAG, and further esterified to TAG. After gavage with 1,3-DAG emulsion, 1,3-DAG was digested mainly to 1(3)-MAG and FFA in the rat small intestinal lumen with subsequent significant increase of 1-MAG and 1,3-DAG concentrations in small intestinal mucosal epithelial cells, and the 2-MAG, 1,2(2,3)-DAG, and TAG concentrations in mucosal epithelial cells were not significantly different after 1,3-DAG than after TAG gavage, suggesting that the metabolic pathway of 1,3-DAG is different from that of TAG. In intestinal mucosal epithelial cells, we further assayed enzyme levels and gene expression of proteins in the phosphatidic acid (PtdOH) pathway. The glycerol kinase, phosphatidate phosphatase, and diacylglycerol acyltransferase-2 expression and the relative expression of mRNA of enzymes were significantly increased in the 1,3-DAG group compared with the TAG group, suggesting that TAG synthesis from dietary 1,3-DAG was mainly via PtdOH pathways, which may partially account for the effect of dietary DAG on postprandial serum TAG.

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Glycerol kinase from Escherichia coli and an Ala65→Thr mutant: the crystal structures reveal conformational changes with implications for allosteric regulation

Cited by 56 publications

References 55 publications

Structure and Function of Human Xylulokinase, an Enzyme with Important Roles in Carbohydrate Metabolism

Structure and Function of Human Xylulokinase, an Enzyme with Important Roles in Carbohydrate Metabolism

Transplanting allosteric control of enzyme activity by protein–protein interactions: Coupling a regulatory site to the conserved catalytic core

Rat Small Intestinal Mucosal Epithelial Cells Absorb Dietary 1,3‐Diacylglycerol Via Phosphatidic Acid Pathways

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