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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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The superimposition of the FFA1 crystal structure and the homology model of the FFA1 wild type bound to TAK-875. The FFA1 wild type model was constructed from the FFA1 crystal structure using Prime [43]. TAK-875 was docked to the wild type model using Glide [20]. Phenylalanine mutation at position 3.34 is shown in pink
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Fig4: The superimposition of the FFA1 crystal structure and the homology model of the FFA1 wild type bound to TAK-875. The FFA1 wild type model was constructed from the FFA1 crystal structure using Prime [43]. TAK-875 was docked to the wild type model using Glide [20]. Phenylalanine mutation at position 3.34 is shown in pink

Mentions: Because the crystal structure of FFA1 is available only in a complex with TAK-875 we next explore docking of a linoleic acid and synthetic agonists in the experimental FFA1 structure. Initially, we validate a docking protocol of the Glide program [20] by re-docking TAK-875 to the crystal structure to examine whether it can reproduce the experimental binding pose of TAK-875. In all generated poses, the Glide standard docking protocol [20] has reproduced the position of TAK-875 close to the crystal structure position with a RMSD value of 0.8 Å. Notably, alanine at position 3.34 in the crystal structure (Fig. 3a) is a mutation being used together with L42A, G103A and Y202F to thermostabilise the protein [16]. The alanine is located ~5 Å from TAK-875, whereas phenylalanine at this position in the wild type could be at distance of 2 Å. Given its potential impact on a ligand position we rebuilt the wild type of FFA1 and docked TAK-875 (Fig. 4).Fig. 4


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

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

The superimposition of the FFA1 crystal structure and the homology model of the FFA1 wild type bound to TAK-875. The FFA1 wild type model was constructed from the FFA1 crystal structure using Prime [43]. TAK-875 was docked to the wild type model using Glide [20]. Phenylalanine mutation at position 3.34 is shown in pink
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4561419&req=5

Fig4: The superimposition of the FFA1 crystal structure and the homology model of the FFA1 wild type bound to TAK-875. The FFA1 wild type model was constructed from the FFA1 crystal structure using Prime [43]. TAK-875 was docked to the wild type model using Glide [20]. Phenylalanine mutation at position 3.34 is shown in pink
Mentions: Because the crystal structure of FFA1 is available only in a complex with TAK-875 we next explore docking of a linoleic acid and synthetic agonists in the experimental FFA1 structure. Initially, we validate a docking protocol of the Glide program [20] by re-docking TAK-875 to the crystal structure to examine whether it can reproduce the experimental binding pose of TAK-875. In all generated poses, the Glide standard docking protocol [20] has reproduced the position of TAK-875 close to the crystal structure position with a RMSD value of 0.8 Å. Notably, alanine at position 3.34 in the crystal structure (Fig. 3a) is a mutation being used together with L42A, G103A and Y202F to thermostabilise the protein [16]. The alanine is located ~5 Å from TAK-875, whereas phenylalanine at this position in the wild type could be at distance of 2 Å. Given its potential impact on a ligand position we rebuilt the wild type of FFA1 and docked TAK-875 (Fig. 4).Fig. 4

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