Molecular reconstruction of recurrent evolutionary switching in olfactory receptor specificity

Olfactory receptor repertoires exhibit remarkable functional diversity, but how these proteins have evolved is poorly understood. Through analysis of extant and ancestrally-reconstructed drosophilid olfactory receptors from the Ionotropic receptor (Ir) family, we investigated evolution of two organic acid-sensing receptors, Ir75a and Ir75b. Despite their low amino acid identity, we identify a common 'hotspot' in their ligand-binding pocket that has a major effect on changing the specificity of both Irs, as well as at least two distinct functional transitions in Ir75a during evolution. Moreover, we show that odor specificity is refined by changes in additional, receptor-specific sites, including those outside the ligand-binding pocket. Our work reveals how a core, common determinant of ligand-tuning acts within epistatic and allosteric networks of substitutions to lead to functional evolution of olfactory receptors.

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Molecular reconstruction of recurrent evolutionary switching in olfactory receptor specificity

10.1101/2021.04.23.440888 ◽

2021 ◽

Author(s):

Lucia L. Prieto-Godino ◽

Hayden R. Schmidt ◽

Richard Benton

Keyword(s):

Ligand Binding ◽

Olfactory Receptor ◽

Olfactory Receptors ◽

Binding Pocket ◽

Amino Acid Identity ◽

Major Effect ◽

Ligand Docking ◽

Ionotropic Receptor ◽

Ligand Binding Pocket ◽

Receptor Interactions

AbstractOlfactory receptor repertoires exhibit remarkable functional diversity, but how these proteins have evolved is poorly understood. Through analysis of extant and ancestrally-reconstructed drosophilid olfactory receptors from the Ionotropic Receptor (IR) family, we investigated evolution of two organic acid-sensing receptors, IR75a and IR75b. Despite their low amino acid identity, we identify a common “hotspot” in their ligand-binding pocket that has a major effect on changing the specificity of both IRs, as well as at least two distinct functional transitions in IR75a during evolution. Ligand-docking into IR models predicts that the hotspot does not contact odor molecules, suggesting that this residue indirectly influences ligand/receptor interactions. Moreover, we show that odor specificity is refined by changes in additional, receptor-specific sites, including those outside the ligand-binding pocket. Our work reveals how a core, common determinant of ligand-tuning acts within epistatic and allosteric networks of substitutions to lead to functional evolution of olfactory receptors.

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Suramin Targets the Conserved Ligand-Binding Pocket of Human Raf1 Kinase Inhibitory Protein

Molecules ◽

10.3390/molecules26041151 ◽

2021 ◽

Vol 26 (4) ◽

pp. 1151

Author(s):

Chenyun Guo ◽

Zhihua Wu ◽

Weiliang Lin ◽

Hao Xu ◽

Ting Chang ◽

...

Keyword(s):

Side Effects ◽

Ligand Binding ◽

Mapk Pathway ◽

Regulatory Protein ◽

Therapeutic Index ◽

Binding Pocket ◽

Biological Macromolecules ◽

Inhibitory Protein ◽

Ligand Binding Pocket ◽

Raf1 Kinase

Suramin was initially used to treat African sleeping sickness and has been clinically tested to treat human cancers and HIV infection in the recent years. However, the therapeutic index is low with numerous clinical side-effects, attributed to its diverse interactions with multiple biological macromolecules. Here, we report a novel binding target of suramin, human Raf1 kinase inhibitory protein (hRKIP), which is an important regulatory protein involved in the Ras/Raf1/MEK/ERK (MAPK) signal pathway. Biolayer interference technology showed that suramin had an intermediate affinity for binding hRKIP with a dissociation constant of 23.8 µM. Both nuclear magnetic resonance technology and molecular docking analysis revealed that suramin bound to the conserved ligand-binding pocket of hRKIP, and that residues K113, W173, and Y181 play crucial roles in hRKIP binding suramin. Furthermore, suramin treatment at 160 µM could profoundly increase the ERK phosphorylation level by around 3 times. Our results indicate that suramin binds to hRKIP and prevents hRKIP from binding with hRaf1, thus promoting the MAPK pathway. This work is beneficial to both mechanistically understanding the side-effects of suramin and efficiently improving the clinical applications of suramin.

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Amino acid divergence in the ligand binding pocket of Vibrio LuxR/HapR proteins determines the efficacy of thiophenesulfonamide inhibitors

Molecular Microbiology ◽

10.1111/mmi.14804 ◽

2021 ◽

Author(s):

Jane D. Newman ◽

Jay Chopra ◽

Priyanka Shah ◽

Eda Shi ◽

Molly E. McFadden ◽

...

Keyword(s):

Amino Acid ◽

Ligand Binding ◽

Binding Pocket ◽

Ligand Binding Pocket ◽

Amino Acid Divergence

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Ligands with poly-fluorophenyl moieties promote a local structural rearrangement in the Spinach2 and Broccoli aptamers that increases ligand affinities

10.1101/2021.10.05.463302 ◽

2021 ◽

Author(s):

Sharif Anisuzzaman ◽

Ivan M Geraskin ◽

Muslum Ilgu ◽

Lee Bendickson ◽

George A Kraus ◽

...

Keyword(s):

Nucleic Acids ◽

Ligand Binding ◽

Small Molecule ◽

Binding Pocket ◽

Structural Rearrangement ◽

Structural Reorganization ◽

Ligand Binding Pocket ◽

Rna Remodeling ◽

Small Molecule Ligands ◽

Target Molecules

The interaction of nucleic acids with their molecular targets often involves structural reorganization that may traverse a complex folding landscape. With the more recent recognition that many RNAs, both coding and noncoding, may regulate cellular activities by interacting with target molecules, it becomes increasingly important to understand the means by which nucleic acids interact with their targets and how drugs might be developed that can influence critical folding transitions. We have extensively investigated the interaction of the Spinach2 and Broccoli aptamers with a library of small molecule ligands modified by various extensions from the imido nitrogen of DFHBI (3,5-difluoro-4-hydroxybenzylidene imidazolinone) that reach out from the Spinach2 ligand binding pocket. Studies of the interaction of these compounds with the aptamers revealed that poly-fluorophenyl-modified ligands initiate a slow change in aptamer affinity that takes an extended time (half-life of ~40 min) to achieve. The change in affinity appears to involve an initial disruption of the entrance to the ligand binding pocket followed by a gradual lockdown for which the most likely driving force is an interaction of the gateway adenine with a nearby 2'OH group. These results suggest that poly-fluorophenyl modifications might increase the ability of small molecule drugs to disrupt local structure and promote RNA remodeling.

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Activation of transcription through the ligand-binding pocket of the orphan nuclear receptor ultraspiracle

European Journal of Biochemistry ◽

10.1046/j.1432-1033.2002.03293.x ◽

2002 ◽

Vol 269 (24) ◽

pp. 6026-6036 ◽

Cited By ~ 60

Author(s):

Yong Xu ◽

Fang Fang ◽

YanXia Chu ◽

Davy Jones ◽

Grace Jones

Keyword(s):

Ligand Binding ◽

Nuclear Receptor ◽

Binding Pocket ◽

Orphan Nuclear Receptor ◽

Ligand Binding Pocket ◽

Activation Of Transcription

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Ligand binding pocket function of Drosophila USP is necessary for metamorphosis

General and Comparative Endocrinology ◽

10.1016/j.ygcen.2012.11.009 ◽

2013 ◽

Vol 182 ◽

pp. 73-82 ◽

Cited By ~ 24

Author(s):

Grace Jones ◽

Peter Teal ◽

Vincent C. Henrich ◽

Anna Krzywonos ◽

Agnes Sapa ◽

...

Keyword(s):

Ligand Binding ◽

Binding Pocket ◽

Ligand Binding Pocket

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Agonist Activation of the Angiotensin II Type 1 Receptor Alters the Spatial Proximity of Transmembrane Domain 7 to the Ligand Binding Pocket

Understanding Biology Using Peptides ◽

10.1007/978-0-387-26575-9_144 ◽

2010 ◽

pp. 357-358

Author(s):

Dany Fillion ◽

Gaétan Guillemette ◽

Richard Leduc ◽

Emanuel Escher

Keyword(s):

Angiotensin Ii ◽

Ligand Binding ◽

Transmembrane Domain ◽

Binding Pocket ◽

Spatial Proximity ◽

Ligand Binding Pocket

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Computational redesign of the Escherichia coli ribose-binding protein ligand binding pocket for 1,3-cyclohexanediol and cyclohexanol

Scientific Reports ◽

10.1038/s41598-019-53507-5 ◽

2019 ◽

Vol 9 (1) ◽

Author(s):

Diogo Tavares ◽

Artur Reimer ◽

Shantanu Roy ◽

Aurélie Joublin ◽

Vladimir Sentchilo ◽

...

Keyword(s):

Escherichia Coli ◽

Ligand Binding ◽

Binding Proteins ◽

Binding Pocket ◽

Wild Type ◽

Ligand Binding Pocket ◽

Mutant Proteins ◽

Periplasmic Binding Proteins ◽

Natural Ligands ◽

Ribose Binding Protein

AbstractBacterial periplasmic-binding proteins have been acclaimed as general biosensing platform, but their range of natural ligands is too limited for optimal development of chemical compound detection. Computational redesign of the ligand-binding pocket of periplasmic-binding proteins may yield variants with new properties, but, despite earlier claims, genuine changes of specificity to non-natural ligands have so far not been achieved. In order to better understand the reasons of such limited success, we revisited here the Escherichia coli RbsB ribose-binding protein, aiming to achieve perceptible transition from ribose to structurally related chemical ligands 1,3-cyclohexanediol and cyclohexanol. Combinations of mutations were computationally predicted for nine residues in the RbsB binding pocket, then synthesized and tested in an E. coli reporter chassis. Two million variants were screened in a microcolony-in-bead fluorescence-assisted sorting procedure, which yielded six mutants no longer responsive to ribose but with 1.2–1.5 times induction in presence of 1 mM 1,3-cyclohexanediol, one of which responded to cyclohexanol as well. Isothermal microcalorimetry confirmed 1,3-cyclohexanediol binding, although only two mutant proteins were sufficiently stable upon purification. Circular dichroism spectroscopy indicated discernable structural differences between these two mutant proteins and wild-type RbsB. This and further quantification of periplasmic-space abundance suggested most mutants to be prone to misfolding and/or with defects in translocation compared to wild-type. Our results thus affirm that computational design and library screening can yield RbsB mutants with recognition of non-natural but structurally similar ligands. The inherent arisal of protein instability or misfolding concomitant with designed altered ligand-binding pockets should be overcome by new experimental strategies or by improved future protein design algorithms.

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