scholarly journals Mapping the ρ1GABACReceptor Agonist Binding Pocket

2004 ◽  
Vol 280 (2) ◽  
pp. 1535-1542 ◽  
Author(s):  
Anna Sedelnikova ◽  
Craig D. Smith ◽  
Stanislav O. Zakharkin ◽  
Delores Davis ◽  
David S. Weiss ◽  
...  
Keyword(s):  
Binding Pocket ◽  
Agonist Binding

The importance of interactions with helix 5 in determining the efficacy of β-adrenoceptor ligands

2013 ◽  
Vol 41 (1) ◽  
pp. 159-165 ◽  
Author(s):  
Tony Warne ◽  
Christopher G. Tate
Keyword(s):  
Transmembrane Helix ◽  
Binding Pocket ◽  
Pharmacological Properties ◽  
Agonist Binding ◽  
Inverse Agonists ◽  
Inactive State ◽  
Partial Agonists ◽  
Activated State

Structures of the inactive state of the thermostabilized β1-adrenoceptor have been determined bound to eight different ligands, including full agonists, partial agonists, inverse agonists and biased agonists. Comparison of the structures shows distinct differences within the binding pocket that correlate with the pharmacological properties of the ligands. These data suggest that full agonists stabilize a structure with a contracted binding pocket and a rotamer change of serine (5.46) compared with when antagonists are bound. Inverse agonists may prevent both of these occurrences, whereas partial agonists stabilize a contraction of the binding pocket but not the rotamer change of serine (5.46). It is likely that subtle changes in the interactions between transmembrane helix 5 (H5) and H3/H4 on agonist binding promote the formation of the activated state.

scholarly journals Allosteric coupling from G protein to the agonist-binding pocket in GPCRs

Nature ◽  
2016 ◽  
Vol 535 (7610) ◽  
pp. 182-186 ◽  
Author(s):  
Brian T. DeVree ◽  
Jacob P. Mahoney ◽  
Gisselle A. Vélez-Ruiz ◽  
Soren G. F. Rasmussen ◽  
Adam J. Kuszak ◽  
...  
Keyword(s):  
G Protein ◽  
Binding Pocket ◽  
Agonist Binding ◽  
Allosteric Coupling

Probing the Agonist Binding Pocket in the Nicotinic Acetylcholine Receptor:  A High-Resolution Solid-State NMR Approach

Biochemistry ◽  
1998 ◽  
Vol 37 (30) ◽  
pp. 10854-10859 ◽  
Author(s):  
P. T. F. Williamson ◽  
G. Gröbner ◽  
P. J. R. Spooner ◽  
K. W. Miller ◽  
A. Watts
Keyword(s):  
Solid State ◽  
High Resolution ◽  
Acetylcholine Receptor ◽  
Nicotinic Acetylcholine Receptor ◽  
Solid State Nmr ◽  
Binding Pocket ◽  
Agonist Binding ◽  
Nicotinic Acetylcholine

scholarly journals Photolabelling the urotensin II receptor reveals distinct agonist- and partial-agonist-binding sites

2007 ◽  
Vol 402 (1) ◽  
pp. 51-61 ◽  
Author(s):  
Brian J. Holleran ◽  
Marie-Eve Beaulieu ◽  
Christophe D. Proulx ◽  
Pierre Lavigne ◽  
Emanuel Escher ◽  
...  
Keyword(s):  
Binding Site ◽  
Conformational Changes ◽  
Binding Sites ◽  
Transmembrane Domain ◽  
Partial Agonist ◽  
Binding Pocket ◽  
Activation Mechanism ◽  
Urotensin Ii ◽  
Agonist Binding ◽  
Partial Agonists

The mechanism by which GPCRs (G-protein-coupled receptors) undergo activation is believed to involve conformational changes following agonist binding. We have used photoaffinity labelling to identify domains within GPCRs that make contact with various photoreactive ligands in order to better understand the activation mechanism. Here, a series of four agonist {[Bpa1]U-II (Bpa is p-benzoyl-L-phenylalanine), [Bpa2]U-II, [Bpa3]U-II and [Bpa4]U-II} and three partial agonist {[Bpa1Pen5D-Trp7Orn8]U-II (Pen is penicillamine), [Bpa2Pen5D-Trp7Orn8]U-II and [Pen5Bpa6D-Trp7Orn8]U-II} photoreactive urotensin II (U-II) analogues were used to identify ligand-binding sites on the UT receptor (U-II receptor). All peptides bound the UT receptor expressed in COS-7 cells with high affinity (Kd of 0.3–17.7 nM). Proteolytic mapping and mutational analysis led to the identification of Met288 of the third extracellular loop of the UT receptor as a binding site for all four agonist peptides. Both partial agonists containing the photoreactive group in positions 1 and 2 also cross-linked to Met288. We found that photolabelling with the partial agonist containing the photoreactive group in position 6 led to the detection of transmembrane domain 5 as a binding site for that ligand. Interestingly, this differs from Met184/Met185 of the fourth transmembrane domain that had been identified previously as a contact site for the full agonist [Bpa6]U-II. These results enable us to better map the binding pocket of the UT receptor. Moreover, the data also suggest that, although structurally related agonists or partial agonists may dock in the same general binding pocket, conformational changes induced by various states of activation may result in slight differences in spatial proximity within the cyclic portion of U-II analogues.

scholarly journals A single molecular distance predicts agonist binding energy in nicotinic receptors

2019 ◽  
Vol 151 (4) ◽  
pp. 452-464 ◽  
Author(s):  
Sushree Tripathy ◽  
Wenjun Zheng ◽  
Anthony Auerbach
Keyword(s):  
Binding Energy ◽  
Acetylcholine Receptors ◽  
Structural Parameters ◽  
Aromatic Amino Acids ◽  
Binding Pocket ◽  
Binding Energies ◽  
Agonist Binding ◽  
Response Curves ◽  
Dynamics Simulations ◽  
Experimental Binding Energy

Agonists turn on receptors because they bind more strongly to active (R*) versus resting (R) conformations of their target sites. Here, to explore how agonists activate neuromuscular acetylcholine receptors, we built homology models of R and R* neurotransmitter binding sites, docked ligands to those sites, ran molecular dynamics simulations to relax (“equilibrate”) the structures, measured binding site structural parameters, and correlated them with experimental agonist binding energies. Each binding pocket is a pyramid formed by five aromatic amino acids and covered partially by loop C. We found that in R* versus R, loop C is displaced outward, the pocket is smaller and skewed, the agonist orientation is reversed, and a key nitrogen atom in the agonist is closer to the pocket center (distance dx) and a tryptophan pair but farther from αY190. Of these differences, the change in dx shows the largest correlation with experimental binding energy and provides a good estimate of agonist affinity, efficacy, and efficiency. Indeed, concentration–response curves can be calculated from just dx values. The contraction and twist of the binding pocket upon activation resemble gating rearrangements of the extracellular domain of related receptors at a smaller scale.

scholarly journals Roles of conserved ectodomain cysteines of the rat P2X4 purinoreceptor in agonist binding and channel gating

2010 ◽  
pp. 927-935
Author(s):  
MB Rokic ◽  
V Tvrdoňová ◽  
V Vávra ◽  
M Jindřichová ◽  
T Obšil ◽  
...  
Keyword(s):  
Disulfide Bonds ◽  
Response Pattern ◽  
Transmembrane Domain ◽  
Binding Pocket ◽  
P2x Receptors ◽  
Hek293 Cells ◽  
Receptor Function ◽  
Agonist Binding ◽  
P2x4 Receptor ◽  
Maximum Current

Mammalian P2X receptors contain 10 conserved cysteine residues in their ectodomains, which form five disulfide bonds (SS1-5). Here, we analyzed the relevance of these disulfide pairs in rat P2X4 receptor function by replacing one or both cysteines with alanine or threonine, expressing receptors in HEK293 cells and studying their responsiveness to ATP in the absence and presence of ivermectin, an allostenic modulator of these channels. Response to ATP was not altered when both cysteines forming the SS3 bond (C132-C159) were replaced with threonines. Replacement of SS1 (C116-C165), SS2 (C126-C149) and SS4 (C217-C227), but not SS5 (C261-C270), cysteine pairs with threonines resulted in decreased sensitivity to ATP and faster deactivation times. The maximum current amplitude was reduced in SS2, SS4 and SS5 double mutants and could be partially rescued by ivermectin in SS2 and SS5 double mutants. This response pattern was also observed in numerous single residue mutants, but receptor function was not affected when the 217 cysteine was replaced with threonine or arginine or when the 261 cysteine was replaced with alanine. These results suggest that the SS1, SS2 and SS4 bonds contribute substantially to the structure of the ligand binding pocket, while the SS5 bond located towards the transmembrane domain contributes to receptor gating.

Characterization of Recombinant Human Coagulation Factor XFriuli

1996 ◽  
Vol 75 (02) ◽  
pp. 313-317 ◽  
Author(s):  
D J Kim ◽  
A Girolami ◽  
H L James
Keyword(s):  
Catalytic Domain ◽  
Coagulation Factor ◽  
Molecular Size ◽  
Binding Pocket ◽  
Site Directed Mutagenesis ◽  
Bleeding Tendency ◽  
Factor X ◽  
Amino Terminal ◽  
Normal Plasma ◽  
Plasma Factor

SummaryNaturally occurring plasma factor XFriuli (pFXFr) is marginally activated by both the extrinsic and intrinsic coagulation pathways and has impaired catalytic potential. These studies were initiated to obtain confirmation that this molecule is multi-functionally defective due to the substitution of Ser for Pro at position 343 in the catalytic domain. By the Nelson-Long site-directed mutagenesis procedure a construct of cDNA in pRc/CMV was derived for recombinant factor XFriuli (rFXFr) produced in human embryonic (293) kidney cells. The rFXFr was purified and shown to have a molecular size identical to that of normal plasma factor X (pFX) by gel electrophoretic, and amino-terminal sequencing revealed normal processing cleavages. Using recombinant normal plasma factor X (rFXN) as a reference, the post-translational y-carboxy-glutamic acid (Gla) and (β-hydroxy aspartic acid (β-OH-Asp) content of rFXFr was over 85% and close to 100%, respectively, of expected levels. The specific activities of rFXFr in activation and catalytic assays were the same as those of pFXFr. Molecular modeling suggested the involvement of a new H-bond between the side-chains of Ser-343 and Thr-318 as they occur in anti-parallel (3-pleated sheets near the substrate-binding pocket of pFXFr. These results support the conclusion that the observed mutation in pFXFr is responsible for its dysfunctional activation and catalytic potentials, and that it accounts for the moderate bleeding tendency in the homozygous individuals who possess this variant procoagulant.

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