Cys-Loop Family of Ligand-Gated Ion Channels
Sarah C R Lummis
Outline
Lecture 1:
- The location and role of Cys-loop receptors
o Transmission at the synapse
o Allows fast information transfer
- The structure of Cys-loop receptors
o Pentameric & Modular
- Ligand binding site
o AChBP
o Cation pi interaction
o Other interactions
Lecture 2:
- The pore
o Structure
o Selectivity
- The intracellular domain
o Targeting
o Modulation
o Conductance
- How ligand binding causes channel opening
o Global changes in protein
o Is GLIC a good model protein? (continued in Lecture 3)
Lecture 3:
- What happens when things go wrong
- Understanding Cys-loop receptors allows us to understand what is happening to the
receptor when there are neurological problems, and how best to treat them
o nACh, GABAA, 5-HT3, glycine receptor
Key Themes: gating (or only focus on opening), structures of bacterial LGICs helping
understand mammalian LGICs, compare pore of different LGICs and discuss prospects for
novel drug design
Example Essays
1. How has AChBP contributed to our understanding of Cys-loop receptor function?
2. Describe, using experimental evidence, how neurotransmitters and other ligands bind to
Cys-loop receptors, and how this results in channel opening.
3. How have studies with unnatural amino acids clarified the role of residues in pLGIC, and
what further questions could you answer with them?
4. “Molecular dynamic simulations using structural data provides the only means of fully
understanding the mechanism of action of Cys-loop receptors”. Discuss.
Past Paper Essays
,2018 (Paper 1) Compare and contrast the pores of different pentameric ligand-gated ion
channels and discuss whether these regions can be used as targets for novel drug design.
2016 (Paper 1) Describe studies that have demonstrated how acetylcholine binds in the
nicotinic acetylcholine receptor binding site. How does this result in subsequent channel
opening?
2015 (Paper 1) How do neurotransmitters such as acetylcholine open ion channels?
2014 (Paper 2) How have structures of bacterial ligand-gated ion channels increased our
understanding of Cys-loop receptor structure and function?
2013 (Paper 1) Has the acetylcholine-binding protein provided useful information to
clarify the structure and function of Cys-loop receptors?
2012 (Paper 1) How is binding coupled to gating in Cys-loop ion channels?
2010 (Paper 1) How are nicotinic receptor channels gated?
2009 (Paper 1) How have structural studies advanced our understanding of the function of
Cys-Loop ion channels?
Reading List
(** well worth reading; * prob worth reading; others for further explanation of a specific
topic.)
Cys-Loop Receptors
Akabas et al. Identification of acetylcholine receptor channel-lining residues in the entire M2
segment of the alpha subunit. Neuron. 1994;13:919-27. The first use of SCAM
**Althoff T, et al. X-ray structures of GluCl in apo states reveal a gating mechanism of Cys-loop
receptors. Nature. 2014; 512:333-7. The first Cys-loop receptor structure in both open and
closed states
*Brejc K, et al. Crystal structure of an ACh-binding protein reveals the ligand-binding domain
of nicotinic receptors. Nature 2001;411:269-76. The first crystal structure of a nACh receptor
mimic
Bouzat C. New insights into the structural basis of activation of Cys-loop receptors J Physiol
2012 ;106:23-33. Good review, although too much emphasis on her work
**Cecchini,M Changeux JP. The nicotinic acetylcholine receptor and its prokaryotic
homologues: Structure, conformational transitions & allosteric modulation Neuropharmacol
96 (2015) 137e149 Good review
Corringer PJ, Le Novere N, Changeux JP. Nicotinic receptors at the amino acid level. Annu
Rev Pharmacol Toxicol. 2000;40:431-58 Good but old review
*da Costa CJ, Baenziger JE. Gating of pentameric ligand-gated ion channels: structural
insights and ambiguities. Structure. 2013 21(8):1271-83.
*Dougherty DA.Cys-loop neuroreceptors: structure to the rescue Chem Rev. 2008
;108:1642-53. Lovely easy to read and chatty review, although a bit dated now
Du, J., Lu, W., Wu, S., Cheng, Y. & Gouaux, E. Glycine receptor mechanism elucidated by
electron cryo-microscopy. Nature (2015). 526:224-9.
Eisele, J.l., et al. (1993) Chimaeric nicotinic-serotonergic receptor combines distinct ligand
binding and channel specificities. Nature 366, 479-483 Shows that binding and channel
functions are in distinct domains
, Gunthorpe MJ, Lummis SC. Conversion of the ion selectivity of the 5-HT(3a) receptor from
cationic to anionic reveals a conserved feature of the ligand-gated ion channel superfamily. J
Biol Chem. 2001 ;276:10977-83
*Hassaine G, et al. X-ray structure of the mouse serotonin 5-HT3 receptor. Nature.
2014;512:276-81. One of the 1st vertebrate Cys loop receptor structures – and the only one
with an ICD
Hansen, S. B. et al. Structures of Aplysia AChBP complexes with nicotinic agonists and
antagonists reveal distinctive binding interfaces and conformations. EMBO J. 24, 3635–3646
(2005).
Hibbs, R. E. et al. Structural determinants for interaction of partial agonists with
acetylcholine binding protein and neuronal alpha7 nicotinic acetylcholine receptor. EMBO J.
28, 3040–3051 (2009).
*Hibbs RE, Gouaux E. Principles of activation and permeation in an anion selective Cys-loop
receptor. Nature. 2011; 474:54-60. The first really high resolution Cys-loop receptor
structure, but it is from an invertebrate
Huang X, Chen H, Michelsen K, Schneider S, Shaffer PL. Crystal structure of human glycine
receptor-α3 bound to antagonist strychnine. Nature. 2015 Oct 8;526(7572):277-80. Another
vertebrate Cys-loop structure
Huang, X., Chen, H. & Shaffer, P. L. Crystal Structures of Human GlyRα3 Bound to Ivermectin.
Structure (2017). doi:10.1016/j.str.2017.04.007
Huang, X. et al. Crystal structures of human glycine receptor α3 bound to a novel class of
analgesic potentiators. Nat. Struct. Mol. Biol. 24, 108–113 (2017).
Jadey, S. & Auerbach, A. An integrated catch-and-hold mechanism activates nicotinic
acetylcholine receptors. J. Gen. Physiol. 140, 17–28 (2012).The catch and hold hypothesis
Jaiteh, M., Taly, A. & Hénin, J. Evolution of Pentameric Ligand-Gated Ion Channels: Pro-Loop
Receptors. PLoS ONE 11, e0151934 (2016).
Jansen, M., Bali, M. & Akabas, M. H. Modular design of Cys-loop ligand-gated ion channels:
functional 5-HT3 and GABA rho1 receptors lacking the large cytoplasmic M3M4 loop. J. Gen.
Physiol. 131, 137–146 (2008).
*Kelley SP, Dunlop JI Kirkness EF Lambert JJ Peters JA A cytoplasmic region determines
single-channel conductance in 5-HT3 receptors. Nature 2003; 424, 321-324 The pore does
not do all the work in controlling ion flow
Lynagh T, Pless SA. Principles of agonist recognition in Cys-loop receptors. Front. Physiol.
2014 Apr 24;5:160.
Lynagh, T. & Lynch, J. W. Molecular mechanisms of Cys-loop ion channel receptor
modulation by ivermectin. Front Mol Neurosci 5, 60 (2012).
*Miller PS, et al. Crystal structure of a human GABAA receptor. Nature. 2014; 512:270-5.
Another vertebrate Cys loop receptor structure but not one found physiologically
*Miller PS, Smart TG. Binding, activation and modulation of Cys-loop receptors .Trends
Pharmacol Sci. 2010 Apr;31(4):161-74. Good review but slightly dated
Morales-Perez CL, Noviello CM, Hibbs RE. Nature. 2016 Oct 3. doi: 10.1038/nature19785. X-
ray structure of the human α4β2 nicotinic receptor. At last a nAChR structure.
**Nemecz Á, Prevost MS, Menny A, Corringer PJ. Neuron. 2016;90:452-70. Emerging
Molecular Mechanisms of Signal Transduction in Pentameric Ligand-Gated Ion Channels.
Excellent review but more detail than needed
*Nys M, Kesters D, Ulens C. Structural insights into Cys-loop receptor function and ligand
recognition. Biochem Pharmacol. 2013; 86:1042-53. Nice review although a bit old now
Sarah C R Lummis
Outline
Lecture 1:
- The location and role of Cys-loop receptors
o Transmission at the synapse
o Allows fast information transfer
- The structure of Cys-loop receptors
o Pentameric & Modular
- Ligand binding site
o AChBP
o Cation pi interaction
o Other interactions
Lecture 2:
- The pore
o Structure
o Selectivity
- The intracellular domain
o Targeting
o Modulation
o Conductance
- How ligand binding causes channel opening
o Global changes in protein
o Is GLIC a good model protein? (continued in Lecture 3)
Lecture 3:
- What happens when things go wrong
- Understanding Cys-loop receptors allows us to understand what is happening to the
receptor when there are neurological problems, and how best to treat them
o nACh, GABAA, 5-HT3, glycine receptor
Key Themes: gating (or only focus on opening), structures of bacterial LGICs helping
understand mammalian LGICs, compare pore of different LGICs and discuss prospects for
novel drug design
Example Essays
1. How has AChBP contributed to our understanding of Cys-loop receptor function?
2. Describe, using experimental evidence, how neurotransmitters and other ligands bind to
Cys-loop receptors, and how this results in channel opening.
3. How have studies with unnatural amino acids clarified the role of residues in pLGIC, and
what further questions could you answer with them?
4. “Molecular dynamic simulations using structural data provides the only means of fully
understanding the mechanism of action of Cys-loop receptors”. Discuss.
Past Paper Essays
,2018 (Paper 1) Compare and contrast the pores of different pentameric ligand-gated ion
channels and discuss whether these regions can be used as targets for novel drug design.
2016 (Paper 1) Describe studies that have demonstrated how acetylcholine binds in the
nicotinic acetylcholine receptor binding site. How does this result in subsequent channel
opening?
2015 (Paper 1) How do neurotransmitters such as acetylcholine open ion channels?
2014 (Paper 2) How have structures of bacterial ligand-gated ion channels increased our
understanding of Cys-loop receptor structure and function?
2013 (Paper 1) Has the acetylcholine-binding protein provided useful information to
clarify the structure and function of Cys-loop receptors?
2012 (Paper 1) How is binding coupled to gating in Cys-loop ion channels?
2010 (Paper 1) How are nicotinic receptor channels gated?
2009 (Paper 1) How have structural studies advanced our understanding of the function of
Cys-Loop ion channels?
Reading List
(** well worth reading; * prob worth reading; others for further explanation of a specific
topic.)
Cys-Loop Receptors
Akabas et al. Identification of acetylcholine receptor channel-lining residues in the entire M2
segment of the alpha subunit. Neuron. 1994;13:919-27. The first use of SCAM
**Althoff T, et al. X-ray structures of GluCl in apo states reveal a gating mechanism of Cys-loop
receptors. Nature. 2014; 512:333-7. The first Cys-loop receptor structure in both open and
closed states
*Brejc K, et al. Crystal structure of an ACh-binding protein reveals the ligand-binding domain
of nicotinic receptors. Nature 2001;411:269-76. The first crystal structure of a nACh receptor
mimic
Bouzat C. New insights into the structural basis of activation of Cys-loop receptors J Physiol
2012 ;106:23-33. Good review, although too much emphasis on her work
**Cecchini,M Changeux JP. The nicotinic acetylcholine receptor and its prokaryotic
homologues: Structure, conformational transitions & allosteric modulation Neuropharmacol
96 (2015) 137e149 Good review
Corringer PJ, Le Novere N, Changeux JP. Nicotinic receptors at the amino acid level. Annu
Rev Pharmacol Toxicol. 2000;40:431-58 Good but old review
*da Costa CJ, Baenziger JE. Gating of pentameric ligand-gated ion channels: structural
insights and ambiguities. Structure. 2013 21(8):1271-83.
*Dougherty DA.Cys-loop neuroreceptors: structure to the rescue Chem Rev. 2008
;108:1642-53. Lovely easy to read and chatty review, although a bit dated now
Du, J., Lu, W., Wu, S., Cheng, Y. & Gouaux, E. Glycine receptor mechanism elucidated by
electron cryo-microscopy. Nature (2015). 526:224-9.
Eisele, J.l., et al. (1993) Chimaeric nicotinic-serotonergic receptor combines distinct ligand
binding and channel specificities. Nature 366, 479-483 Shows that binding and channel
functions are in distinct domains
, Gunthorpe MJ, Lummis SC. Conversion of the ion selectivity of the 5-HT(3a) receptor from
cationic to anionic reveals a conserved feature of the ligand-gated ion channel superfamily. J
Biol Chem. 2001 ;276:10977-83
*Hassaine G, et al. X-ray structure of the mouse serotonin 5-HT3 receptor. Nature.
2014;512:276-81. One of the 1st vertebrate Cys loop receptor structures – and the only one
with an ICD
Hansen, S. B. et al. Structures of Aplysia AChBP complexes with nicotinic agonists and
antagonists reveal distinctive binding interfaces and conformations. EMBO J. 24, 3635–3646
(2005).
Hibbs, R. E. et al. Structural determinants for interaction of partial agonists with
acetylcholine binding protein and neuronal alpha7 nicotinic acetylcholine receptor. EMBO J.
28, 3040–3051 (2009).
*Hibbs RE, Gouaux E. Principles of activation and permeation in an anion selective Cys-loop
receptor. Nature. 2011; 474:54-60. The first really high resolution Cys-loop receptor
structure, but it is from an invertebrate
Huang X, Chen H, Michelsen K, Schneider S, Shaffer PL. Crystal structure of human glycine
receptor-α3 bound to antagonist strychnine. Nature. 2015 Oct 8;526(7572):277-80. Another
vertebrate Cys-loop structure
Huang, X., Chen, H. & Shaffer, P. L. Crystal Structures of Human GlyRα3 Bound to Ivermectin.
Structure (2017). doi:10.1016/j.str.2017.04.007
Huang, X. et al. Crystal structures of human glycine receptor α3 bound to a novel class of
analgesic potentiators. Nat. Struct. Mol. Biol. 24, 108–113 (2017).
Jadey, S. & Auerbach, A. An integrated catch-and-hold mechanism activates nicotinic
acetylcholine receptors. J. Gen. Physiol. 140, 17–28 (2012).The catch and hold hypothesis
Jaiteh, M., Taly, A. & Hénin, J. Evolution of Pentameric Ligand-Gated Ion Channels: Pro-Loop
Receptors. PLoS ONE 11, e0151934 (2016).
Jansen, M., Bali, M. & Akabas, M. H. Modular design of Cys-loop ligand-gated ion channels:
functional 5-HT3 and GABA rho1 receptors lacking the large cytoplasmic M3M4 loop. J. Gen.
Physiol. 131, 137–146 (2008).
*Kelley SP, Dunlop JI Kirkness EF Lambert JJ Peters JA A cytoplasmic region determines
single-channel conductance in 5-HT3 receptors. Nature 2003; 424, 321-324 The pore does
not do all the work in controlling ion flow
Lynagh T, Pless SA. Principles of agonist recognition in Cys-loop receptors. Front. Physiol.
2014 Apr 24;5:160.
Lynagh, T. & Lynch, J. W. Molecular mechanisms of Cys-loop ion channel receptor
modulation by ivermectin. Front Mol Neurosci 5, 60 (2012).
*Miller PS, et al. Crystal structure of a human GABAA receptor. Nature. 2014; 512:270-5.
Another vertebrate Cys loop receptor structure but not one found physiologically
*Miller PS, Smart TG. Binding, activation and modulation of Cys-loop receptors .Trends
Pharmacol Sci. 2010 Apr;31(4):161-74. Good review but slightly dated
Morales-Perez CL, Noviello CM, Hibbs RE. Nature. 2016 Oct 3. doi: 10.1038/nature19785. X-
ray structure of the human α4β2 nicotinic receptor. At last a nAChR structure.
**Nemecz Á, Prevost MS, Menny A, Corringer PJ. Neuron. 2016;90:452-70. Emerging
Molecular Mechanisms of Signal Transduction in Pentameric Ligand-Gated Ion Channels.
Excellent review but more detail than needed
*Nys M, Kesters D, Ulens C. Structural insights into Cys-loop receptor function and ligand
recognition. Biochem Pharmacol. 2013; 86:1042-53. Nice review although a bit old now
