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Loop B is a major structural component of the 5-HT3 receptor.

Thompson AJ, Lochner M, Lummis SC - Biophys. J. (2008)

Bottom Line: Homology modeling indicates that loop B contributes two residues to a hydrophobic core that faces into the beta-sandwich of the subunit, and the experimental data indicate that they are important for both the structure and the function of the receptor.The models also show that close to the apex of the loop (Ser-182 to Ile-190), loop B residues form an extensive network of hydrogen bonds, both with other loop B residues and with adjacent regions of the protein.Overall, the data suggest that loop B has a major role in maintaining the structure of the region by a series of noncovalent interactions that are easily disrupted by amino acid substitutions.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Cambridge, Cambridge, CB2 1QW, United Kingdom.

ABSTRACT
The 5-HT(3) receptor belongs to a family of therapeutically important neurotransmitter-gated receptors whose ligand binding sites are formed by the convergence of six peptide loops (A-F). Here we have mutated 15 amino acid residues in and around loop B of the 5-HT(3) receptor (Ser-177 to Asn-191) to Ala or a residue with similar chemical properties. Changes in [3H]granisetron binding affinity (K(d)) and 5-HT EC(50) were determined using receptors expressed in human embryonic kidney 293 cells. Substitutions at all but one residue (Thr-181) altered or eliminated binding for one or both mutants. Receptors were nonfunctional or EC(50) values were altered for all but two mutants (S182T, I190L). Homology modeling indicates that loop B contributes two residues to a hydrophobic core that faces into the beta-sandwich of the subunit, and the experimental data indicate that they are important for both the structure and the function of the receptor. The models also show that close to the apex of the loop (Ser-182 to Ile-190), loop B residues form an extensive network of hydrogen bonds, both with other loop B residues and with adjacent regions of the protein. Overall, the data suggest that loop B has a major role in maintaining the structure of the region by a series of noncovalent interactions that are easily disrupted by amino acid substitutions.

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Homology models showing the amino acid backbone of the loop B region of the 5-HT3 receptor. (A) Unbound structure (modeled from AChBP with no ligand, PDB ID 2byn). (B) Agonist bound, (modeled from AChBP with carbomylcholine, PDB ID 1uv6). (C) Agonist bound, (modeled from AChBP with epibatidine, PDB ID 2byq). (D) Large antagonist bound (modeled from AChBP with α-cobratoxin, PDB ID 1yi5). (E) Small antagonist bound, (modeled from AChBP with 2-methyllcaconitine, PDB ID 2byr) (F) Overlays of the modeled loop B backbones from A–E compiled with Swiss-PdbViewer “magic fit”, using residues Ser-177 to Asn-191 as a reference point.
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fig4: Homology models showing the amino acid backbone of the loop B region of the 5-HT3 receptor. (A) Unbound structure (modeled from AChBP with no ligand, PDB ID 2byn). (B) Agonist bound, (modeled from AChBP with carbomylcholine, PDB ID 1uv6). (C) Agonist bound, (modeled from AChBP with epibatidine, PDB ID 2byq). (D) Large antagonist bound (modeled from AChBP with α-cobratoxin, PDB ID 1yi5). (E) Small antagonist bound, (modeled from AChBP with 2-methyllcaconitine, PDB ID 2byr) (F) Overlays of the modeled loop B backbones from A–E compiled with Swiss-PdbViewer “magic fit”, using residues Ser-177 to Asn-191 as a reference point.

Mentions: To gain further insight into the possible orientations and interactions of loop B residues, a range of homology models were constructed based on AChBP crystal structures that were either unbound or bound by different ligands. The lowest energy states for the models are shown in Fig. 4, A–E, and the carbon backbones for each of these are overlaid in Fig. 4 F. Loop B is similar in each of these models, suggesting that regardless of the bound ligand, the position of the loop and the orientations of the side-chains remains relatively unaltered.


Loop B is a major structural component of the 5-HT3 receptor.

Thompson AJ, Lochner M, Lummis SC - Biophys. J. (2008)

Homology models showing the amino acid backbone of the loop B region of the 5-HT3 receptor. (A) Unbound structure (modeled from AChBP with no ligand, PDB ID 2byn). (B) Agonist bound, (modeled from AChBP with carbomylcholine, PDB ID 1uv6). (C) Agonist bound, (modeled from AChBP with epibatidine, PDB ID 2byq). (D) Large antagonist bound (modeled from AChBP with α-cobratoxin, PDB ID 1yi5). (E) Small antagonist bound, (modeled from AChBP with 2-methyllcaconitine, PDB ID 2byr) (F) Overlays of the modeled loop B backbones from A–E compiled with Swiss-PdbViewer “magic fit”, using residues Ser-177 to Asn-191 as a reference point.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2599823&req=5

fig4: Homology models showing the amino acid backbone of the loop B region of the 5-HT3 receptor. (A) Unbound structure (modeled from AChBP with no ligand, PDB ID 2byn). (B) Agonist bound, (modeled from AChBP with carbomylcholine, PDB ID 1uv6). (C) Agonist bound, (modeled from AChBP with epibatidine, PDB ID 2byq). (D) Large antagonist bound (modeled from AChBP with α-cobratoxin, PDB ID 1yi5). (E) Small antagonist bound, (modeled from AChBP with 2-methyllcaconitine, PDB ID 2byr) (F) Overlays of the modeled loop B backbones from A–E compiled with Swiss-PdbViewer “magic fit”, using residues Ser-177 to Asn-191 as a reference point.
Mentions: To gain further insight into the possible orientations and interactions of loop B residues, a range of homology models were constructed based on AChBP crystal structures that were either unbound or bound by different ligands. The lowest energy states for the models are shown in Fig. 4, A–E, and the carbon backbones for each of these are overlaid in Fig. 4 F. Loop B is similar in each of these models, suggesting that regardless of the bound ligand, the position of the loop and the orientations of the side-chains remains relatively unaltered.

Bottom Line: Homology modeling indicates that loop B contributes two residues to a hydrophobic core that faces into the beta-sandwich of the subunit, and the experimental data indicate that they are important for both the structure and the function of the receptor.The models also show that close to the apex of the loop (Ser-182 to Ile-190), loop B residues form an extensive network of hydrogen bonds, both with other loop B residues and with adjacent regions of the protein.Overall, the data suggest that loop B has a major role in maintaining the structure of the region by a series of noncovalent interactions that are easily disrupted by amino acid substitutions.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Cambridge, Cambridge, CB2 1QW, United Kingdom.

ABSTRACT
The 5-HT(3) receptor belongs to a family of therapeutically important neurotransmitter-gated receptors whose ligand binding sites are formed by the convergence of six peptide loops (A-F). Here we have mutated 15 amino acid residues in and around loop B of the 5-HT(3) receptor (Ser-177 to Asn-191) to Ala or a residue with similar chemical properties. Changes in [3H]granisetron binding affinity (K(d)) and 5-HT EC(50) were determined using receptors expressed in human embryonic kidney 293 cells. Substitutions at all but one residue (Thr-181) altered or eliminated binding for one or both mutants. Receptors were nonfunctional or EC(50) values were altered for all but two mutants (S182T, I190L). Homology modeling indicates that loop B contributes two residues to a hydrophobic core that faces into the beta-sandwich of the subunit, and the experimental data indicate that they are important for both the structure and the function of the receptor. The models also show that close to the apex of the loop (Ser-182 to Ile-190), loop B residues form an extensive network of hydrogen bonds, both with other loop B residues and with adjacent regions of the protein. Overall, the data suggest that loop B has a major role in maintaining the structure of the region by a series of noncovalent interactions that are easily disrupted by amino acid substitutions.

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