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Free fatty acid receptors: structural models and elucidation of ligand binding interactions.

Tikhonova IG, Poerio E - BMC Struct. Biol. (2015)

Bottom Line: This binding mode can explain mutagenesis results for residues at positions 4.56 and 5.42.The novel structural models of FFAs provide information on specific modes of ligand binding at FFA subtypes and new suggestions for mutagenesis and ligand modification, guiding the development of novel orthosteric and allosteric chemical probes to validate the importance of FFAs in metabolic and inflammatory conditions.Using our FFA homology modelling experience, a strategy to model a GPCR, which is phylogenetically distant from GPCRs with the available crystal structures, is discussed.

View Article: PubMed Central - PubMed

Affiliation: Molecular Therapeutics, School of Pharmacy, Medical Biology Centre, Queen's University Belfast, Belfast, BT9 7BL, Northern Ireland, UK. i.tikhonova@qub.ac.uk.

ABSTRACT

Background: The free fatty acid receptors (FFAs), including FFA1 (orphan name: GPR40), FFA2 (GPR43) and FFA3 (GPR41) are G protein-coupled receptors (GPCRs) involved in energy and metabolic homeostasis. Understanding the structural basis of ligand binding at FFAs is an essential step toward designing potent and selective small molecule modulators.

Results: We analyse earlier homology models of FFAs in light of the newly published FFA1 crystal structure co-crystallized with TAK-875, an ago-allosteric ligand, focusing on the architecture of the extracellular binding cavity and agonist-receptor interactions. The previous low-resolution homology models of FFAs were helpful in highlighting the location of the ligand binding site and the key residues for ligand anchoring. However, homology models were not accurate in establishing the nature of all ligand-receptor contacts and the precise ligand-binding mode. From analysis of structural models and mutagenesis, it appears that the position of helices 3, 4 and 5 is crucial in ligand docking. The FFA1-based homology models of FFA2 and FFA3 were constructed and used to compare the FFA subtypes. From docking studies we propose an alternative binding mode for orthosteric agonists at FFA1 and FFA2, involving the interhelical space between helices 4 and 5. This binding mode can explain mutagenesis results for residues at positions 4.56 and 5.42. The novel FFAs structural models highlight higher aromaticity of the FFA2 binding cavity and higher hydrophilicity of the FFA3 binding cavity. The role of the residues at the second extracellular loop used in mutagenesis is reanalysed. The third positively-charged residue in the binding cavity of FFAs, located in helix 2, is identified and predicted to coordinate allosteric modulators.

Conclusions: The novel structural models of FFAs provide information on specific modes of ligand binding at FFA subtypes and new suggestions for mutagenesis and ligand modification, guiding the development of novel orthosteric and allosteric chemical probes to validate the importance of FFAs in metabolic and inflammatory conditions. Using our FFA homology modelling experience, a strategy to model a GPCR, which is phylogenetically distant from GPCRs with the available crystal structures, is discussed.

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Ligand binding in FFA2 and FFA3. a: the binding mode of tiglic acid at FFA2. b: the binding mode of 1-MCPC in FFA3. c: the binding mode of 1 in FFA2. FFA2-3 homology models are based on the FFA1 crystal structure. H-bonding and π- π interactions are shown in black and blue dotted lines, respectively
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Fig7: Ligand binding in FFA2 and FFA3. a: the binding mode of tiglic acid at FFA2. b: the binding mode of 1-MCPC in FFA3. c: the binding mode of 1 in FFA2. FFA2-3 homology models are based on the FFA1 crystal structure. H-bonding and π- π interactions are shown in black and blue dotted lines, respectively

Mentions: Docking of the selective short carboxylic acids [10], FFA2-selective tiglic acid and FFA3-selective 1-MCPC to the FFA1-based model of FFA2 and FFA3 indicates that the carboxyl group of the fatty acids is anchored by conserved R1805.39, R2557.35, Y943.37 and Y2386.51. The non-conserved residues at positions 5.43 and E166/L171EL2 predicted to be important for FFA2/FFA3 selectivity together with the residue at position 5.42 in the earlier homology models contribute more to the shape of the binding cavity rather than direct interactions with a ligand (Fig. 7a and b). Instead, the non-conserved residue at position 5.42 is at distance of 4 Å enabling it to form van der Waals interactions. The hydrophobic part of the tiglic acid forms hydrophobic contacts with Y903.33, Y943.37, C1414.57, I1454.61, V1795.38 and Y165EL2 in FFA2, whereas 1-MCPC interacts with F963.33, L1815.35, V1845.38, R1855.39, M1885.42 and F173EL2 in FFA3. The new structural models suggest a small binding cavity in FFA2 due to bulky aromatic residues and an intensive H-bonding network, structurally explaining a preference in binding of carboxylic acids with sp2 hybridized α-carbons to FFA2, whereas a larger binding cavity in FFA3 with a lesser network of interactions could be a reason for a preference in binding of carboxylic acids with sp3 hybridized α-carbons to FFA3 as observed in pharmacological studies [10]. We predict that the residues that determine subtype selectivity between FFA2 and FFA3 are Y903.33, I1454.61 and E166EL2 in FFA2 and the corresponded F963.33, Y1514.61 and L171EL2 in FFA3.Fig. 7


Free fatty acid receptors: structural models and elucidation of ligand binding interactions.

Tikhonova IG, Poerio E - BMC Struct. Biol. (2015)

Ligand binding in FFA2 and FFA3. a: the binding mode of tiglic acid at FFA2. b: the binding mode of 1-MCPC in FFA3. c: the binding mode of 1 in FFA2. FFA2-3 homology models are based on the FFA1 crystal structure. H-bonding and π- π interactions are shown in black and blue dotted lines, respectively
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4561419&req=5

Fig7: Ligand binding in FFA2 and FFA3. a: the binding mode of tiglic acid at FFA2. b: the binding mode of 1-MCPC in FFA3. c: the binding mode of 1 in FFA2. FFA2-3 homology models are based on the FFA1 crystal structure. H-bonding and π- π interactions are shown in black and blue dotted lines, respectively
Mentions: Docking of the selective short carboxylic acids [10], FFA2-selective tiglic acid and FFA3-selective 1-MCPC to the FFA1-based model of FFA2 and FFA3 indicates that the carboxyl group of the fatty acids is anchored by conserved R1805.39, R2557.35, Y943.37 and Y2386.51. The non-conserved residues at positions 5.43 and E166/L171EL2 predicted to be important for FFA2/FFA3 selectivity together with the residue at position 5.42 in the earlier homology models contribute more to the shape of the binding cavity rather than direct interactions with a ligand (Fig. 7a and b). Instead, the non-conserved residue at position 5.42 is at distance of 4 Å enabling it to form van der Waals interactions. The hydrophobic part of the tiglic acid forms hydrophobic contacts with Y903.33, Y943.37, C1414.57, I1454.61, V1795.38 and Y165EL2 in FFA2, whereas 1-MCPC interacts with F963.33, L1815.35, V1845.38, R1855.39, M1885.42 and F173EL2 in FFA3. The new structural models suggest a small binding cavity in FFA2 due to bulky aromatic residues and an intensive H-bonding network, structurally explaining a preference in binding of carboxylic acids with sp2 hybridized α-carbons to FFA2, whereas a larger binding cavity in FFA3 with a lesser network of interactions could be a reason for a preference in binding of carboxylic acids with sp3 hybridized α-carbons to FFA3 as observed in pharmacological studies [10]. We predict that the residues that determine subtype selectivity between FFA2 and FFA3 are Y903.33, I1454.61 and E166EL2 in FFA2 and the corresponded F963.33, Y1514.61 and L171EL2 in FFA3.Fig. 7

Bottom Line: This binding mode can explain mutagenesis results for residues at positions 4.56 and 5.42.The novel structural models of FFAs provide information on specific modes of ligand binding at FFA subtypes and new suggestions for mutagenesis and ligand modification, guiding the development of novel orthosteric and allosteric chemical probes to validate the importance of FFAs in metabolic and inflammatory conditions.Using our FFA homology modelling experience, a strategy to model a GPCR, which is phylogenetically distant from GPCRs with the available crystal structures, is discussed.

View Article: PubMed Central - PubMed

Affiliation: Molecular Therapeutics, School of Pharmacy, Medical Biology Centre, Queen's University Belfast, Belfast, BT9 7BL, Northern Ireland, UK. i.tikhonova@qub.ac.uk.

ABSTRACT

Background: The free fatty acid receptors (FFAs), including FFA1 (orphan name: GPR40), FFA2 (GPR43) and FFA3 (GPR41) are G protein-coupled receptors (GPCRs) involved in energy and metabolic homeostasis. Understanding the structural basis of ligand binding at FFAs is an essential step toward designing potent and selective small molecule modulators.

Results: We analyse earlier homology models of FFAs in light of the newly published FFA1 crystal structure co-crystallized with TAK-875, an ago-allosteric ligand, focusing on the architecture of the extracellular binding cavity and agonist-receptor interactions. The previous low-resolution homology models of FFAs were helpful in highlighting the location of the ligand binding site and the key residues for ligand anchoring. However, homology models were not accurate in establishing the nature of all ligand-receptor contacts and the precise ligand-binding mode. From analysis of structural models and mutagenesis, it appears that the position of helices 3, 4 and 5 is crucial in ligand docking. The FFA1-based homology models of FFA2 and FFA3 were constructed and used to compare the FFA subtypes. From docking studies we propose an alternative binding mode for orthosteric agonists at FFA1 and FFA2, involving the interhelical space between helices 4 and 5. This binding mode can explain mutagenesis results for residues at positions 4.56 and 5.42. The novel FFAs structural models highlight higher aromaticity of the FFA2 binding cavity and higher hydrophilicity of the FFA3 binding cavity. The role of the residues at the second extracellular loop used in mutagenesis is reanalysed. The third positively-charged residue in the binding cavity of FFAs, located in helix 2, is identified and predicted to coordinate allosteric modulators.

Conclusions: The novel structural models of FFAs provide information on specific modes of ligand binding at FFA subtypes and new suggestions for mutagenesis and ligand modification, guiding the development of novel orthosteric and allosteric chemical probes to validate the importance of FFAs in metabolic and inflammatory conditions. Using our FFA homology modelling experience, a strategy to model a GPCR, which is phylogenetically distant from GPCRs with the available crystal structures, is discussed.

Show MeSH
Related in: MedlinePlus