Structural Determinants of Benzodiazepine Allosteric Regulation of GABAA Receptor Currents

Abstract Structure-function studies contribute to deciphering how small modifications in the primary structure could introduce desirable characteristics into enzymes without affecting its overall functioning. Malic enzymes (ME) are ubiquitous and responsible for a wide variety of functions. The availability of a high number of ME crystal structures from different species facilitates comparisons between sequence and structure. Specifically, the structural determinants necessary for fumarate allosteric regulation of ME has been of particular interest. NADP-ME2 from Arabidopsis thaliana exhibits a distinctive and complex regulation by fumarate, acting as an activator or an inhibitor according to substrate and effector concentrations. However, the 3D structure for this enzyme is not yet reported. In this work, we characterized the NADP-ME2 allosteric site by structural modeling, molecular docking, normal mode analysis and mutagenesis. The regulatory site model and its docking analysis suggested that other C4 acids including malate, NADP-ME2 substrate, could also fit into fumarate’s pocket. Besides, a non-conserved cluster of hydrophobic residues in the second sphere of the allosteric site was identified. The substitution of one of those residues, L62, by a less flexible residue as tryptophan, resulted in a complete loss of fumarate activation and a reduction of substrate affinities for the active site. In addition, normal mode analysis indicated that conformational changes leading to the activation could originate in the region surrounding L62, extending through the allosteric site till the active site. Finally, the results in this work contribute to the understanding of structural determinants necessary for allosteric regulation providing new insights for enzyme optimization.

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Structural determinants of allosteric regulation in alternatively spliced AMPA receptors

Neuron ◽

10.1016/0896-6273(95)90227-9 ◽

1995 ◽

Vol 14 (4) ◽

pp. 833-843 ◽

Cited By ~ 105

Author(s):

Kathryn M Partin ◽

Derek Bowie ◽

Mark L Mayer

Keyword(s):

Ampa Receptors ◽

Allosteric Regulation ◽

Structural Determinants ◽

Alternatively Spliced

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Allosteric Regulation of the Photosynthetic C4 Isoenzyme of Phosphoenolpyruvate Carboxylase: A Comparative Study Between Enzymes from Monocot and Dicot Plants

Journal of the Mexican Chemical Society ◽

10.29356/jmcs.v56i1.275 ◽

2017 ◽

Vol 56 (1) ◽

Author(s):

Rodrigo Güémez-Toro ◽

Carlos Mújica-Jiménez ◽

Rosario A. Muñoz-Clares

Keyword(s):

Phosphoenolpyruvate Carboxylase ◽

Allosteric Regulation ◽

Three Dimensional ◽

Dimensional Structure ◽

Velocity Data ◽

Structural Determinants ◽

Multiple Alignments ◽

High Flexibility ◽

Loop Conformation ◽

Conserved Residue

In the present study, we have investigated the complex allosteric regulation of the non-phosphorylated forms of the photosynthetic phosphoenolpyruvate carboxylase isoenzymes (PEPC-C4) from amaranth (AhPEPC-C4) and maize (ZmPEPC-C4) leaves. Previous studies showed that glycine (Gly) only activates PEPC-C4 from monocot plants, as maize, but not from dicot plants, as amaranth. Our initial velocity data confirm this, in spite that AhPEPC-C4 binds Gly with much higher affinity than ZmPEPC-C4. In AhPEPC-C4, the lack of Gly activation is overcome mainly by its higher affinity for the substrate phosphoenolpyruvate and its lower affinity for the inhibitor malate compared with ZmPEPC-C4. We have also explored the structural determinants of the differences in Gly activation by performing multiple alignments between the known monocot and dicot PEPC-C4 sequences and by modeling, in both the AhPEPC-C4 and ZmPEPC-C4 isoenzymes, the three-dimensional structure of the loop proposed as the Gly binding site, which was not observed in the crystal structure of the maize enzyme due to its high flexibility. The models suggest that conserved lysyl and aspartyl residues are important for binding to the activator molecule, and that a nearby non-conserved residue may be responsible for differences between the amaranth and maize enzymes in the loop conformation, which would account for the poorer affinity for Gly of the maize enzyme as well as for its higher degree of activation.

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259: Melatonin Enlarges the Bladder by the Gabaa Receptor in the Brain, Suggesting its Efficaciousness Against Nocturia

The Journal of Urology ◽

10.1016/s0022-5347(18)30524-x ◽

2007 ◽

Vol 177 (4S) ◽

pp. 86-87

Author(s):

Yosuke Matsuta ◽

Aniwar Yusup ◽

Masaharu Nakai ◽

Kazuya Tanase ◽

Yoshitaka Aoki ◽

...

Keyword(s):

Gabaa Receptor ◽

The Brain

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Allosteric regulation of GPCR Rhodopsin by soft matter

10.1021/scimeetings.0c06874 ◽

2020 ◽

Author(s):

Nipuna Weerasinghe ◽

Steven Fried ◽

Anna Eitel ◽

Andrey Struts ◽

Suchithranga Perera ◽

...

Keyword(s):

Soft Matter ◽

Allosteric Regulation

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Cinnamamides from Piper capense with affinity to the benzodiazepine site on the GABAA receptor

Planta Medica ◽

10.1055/s-0028-1084787 ◽

2008 ◽

Vol 74 (09) ◽

Author(s):

ME Pedersen ◽

HB Rasmussen ◽

B Metzler ◽

GI Stafford ◽

J van Staden ◽

...

Keyword(s):

Gabaa Receptor ◽

Benzodiazepine Site

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Bioactivity-guided Isolation of GABAA Receptor Modulating Constituents from the Rhizome of Actaea racemosa

Planta Medica ◽

10.1055/s-0031-1282155 ◽

2011 ◽

Vol 77 (12) ◽

Author(s):

SS Cicek ◽

S Khom ◽

B Taferner ◽

S Hering ◽

H Stuppner

Keyword(s):

Gabaa Receptor ◽

Actaea Racemosa

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Isoflavonoid derivatives from Adenocarpus cincinnatus as positive GABAA receptor modulators

Planta Medica ◽

10.1055/s-0033-1352179 ◽

2013 ◽

Vol 79 (13) ◽

Author(s):

DC Rueda ◽

M de Mieri ◽

S Hering ◽

M Hamburger

Keyword(s):

Gabaa Receptor ◽

Receptor Modulators

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Induction of Endothelial Tissue Factor by Endotoxin and Its Precursors

Thrombosis and Haemostasis ◽

10.1055/s-0038-1649653 ◽

1993 ◽

Vol 70 (04) ◽

pp. 702-706 ◽

Cited By ~ 16

Author(s):

Charles F Moldow ◽

Ronald R Bach ◽

Katherine Staskus ◽

Paul D Rick

Keyword(s):

Tissue Factor ◽

Lipid A ◽

Umbilical Vein ◽

Structural Determinants ◽

Monophosphoryl Lipid A ◽

Maximal Induction ◽

Umbilical Vein Endothelial Cells ◽

Acyloxyacyl Hydrolase ◽

Endothelial Tissue ◽

Specific Mrna

SummaryThe structural determinants of lipopolysaccharide required for the induction of tissue factor in human umbilical vein endothelial cells were studied. Intact lipid A was essential for the induction of tissue factor whereas the incomplete lipid A precursors lipid IVA and lipid X, as well as monophosphoryl lipid A and acyloxyacyl hydrolase-treated lipopolysaccharide, were unable to induce tissue factor and tissue factor specific mRNA. However, the lipid A precursor, lipid IVA, was able to inhibit LPS-mediated induction of tissue factor; structural determinants distal to lipid A were found to be required for maximal induction of tissue factor activity and tissue factor mRNA. The presence of serum in the assay was found to amplify but was not obligate for tissue factor induction by LPS.

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