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Human metabotropic glutamate receptor 5 transmembrane domain

Human metabotropic glutamate receptor 5 transmembrane domain bound to mavoglurant (PDB code of 4oo9). The 7 helices comprise the bulk of the protein structure. mGlu5 receptor is an important part of the glutamate signaling pathway

Drag the structure with the mouse to rotate

Receiving and responding to extracellular messages is critical to the proper function of the nervous system. Glutamate is the primary excitory neurotransmitter of the Central Nervous System (CNS), and metabotropic glutamate receptor 5 is a key member of the glutamate signaling pathway^[1]. Metabotropic glutamate receptor 5 is a homodimeric GPCR that resides in the cellular membrane ^[1]. mGlu5 is a member of the Class C GPCR family and can further be categorized into the Group I subgroup^[2].

The functionality of the mGlu5 receptor is determined by conformational changes throughout multiple domains. mGlu5 will bind glutamate through its extracellular Venus flytrap domain and the signal will be transduced across the membrane to a heterotrimeric G protein, which will ultimately lead to calcium release and the activation of PKC^[2]. The signal is relayed through a Gq/11 pathway^[1]. Activated PKC will elicit a excitory post-synaptic response and modulate long term potentiation in the CNS^[2].

Human mGlu5 is found throughout the central nervous system. Areas containing high concentrations of mGlu5 are often involved in emotional processing and higher cognition^[3]. The localization of mGlu5 in the CNS and the presence of multiple domains makes mGlu5 an excellent target for treating neurological conditions including: schizophrenia, Fragile X, [1], anxiety, and Alzheimer's disease^[2].

1 Discovery
2 Structure
3 Disease
- 3.1 Fragile X
4 References
5 External Resources

Discovery

The mGlu family of receptors was the first of the Class C GPCR to be extensively studied^[2]. The first regions of the protein crystallized and studied were the Venus Fly Trap domain and the cysteine -rich domain (CRD) on the extracellular region of the receptor^[1].The Venus Fly Trap domain is a large extracellular domain that will selectively bind to glutamate^[2]. The CRD is a somewhat smaller domain composed of many ß sheets and cysteine residues ^[2]. The CRD acts as a signal mediator between the Venus Flytrap domain and the transmembrane domain (TMD) of mGlu5, by linking to each domain with disulfide bonds^[2]. The hydrophobic nature and flexibility of the mGlu5 TMD made it difficult to crystallize.

Recently, the human metabotropic glutamate receptor 5 transmembrane domain was crystallized and a structure elucidated^[1]. Several modifications were made to the TMD for successful crystallization. The protein was thermostabilized and flexible domains were removed^[1]. In total residues 2-568 and 837-1153 were excised from the structure. These flexible domains allow mGlu5 to bind to its GPCR ^[1]. The structure of the alpha helices are shown in green, and the negative allosteric modulator mavoglurant shown in yellow. Also in orange, a T4 - was inserted into intracellular loop 2 to add stability^[1].

Structure

Figure 1: Overall Structure of the mGlu5 TMD. The polar heads on the Oleic acids orient the image with the top oriented extracellularly, the middle portion inserted into the membrane, and the lower portion oriented intracellularly. The white exterior represents the surface of the protien, and the multicolored lines interior to the surface represent the backbones 7 transmembrane alpha helices.

Overview

The mGlu5 TMD contains 7 that spans the membrane. The protein was crystallized with shown in red. On the top portion of the protein several critical extracellular loops connect the TMD to the CRD. The binding pocket can be found deep in the interior of the protein, and is mainly comprised of hydrophobic amino acids with more polar amino acids found in the upper and lower portions of the binding site^[1]. Inserted into the biding pocket is the negative allosteric modulator mavoglurant. The TMD is in an inactive conformation, since mavoglurant is bound^[1]. Also, the deletion of the flexible domains leaves the mGlu5 receptor unable to bind to its GPCR^[1]. The inactive state is maintained by multiple ionic locks whose positions determine the active versus inactive conformation.

When mGlu5 is in the active conformation, signaling begins with glutamate binding to the Venus flytrap domain. The signal is transduced across the cysteine-rich domain to the TMD^[3]. Next, the dimerization of the TMD occurs. This activates the Gq/11 pathway, which activates phospholipase Cβ^[3]. The active phospholipase Cβ hydrolyzes phosphotinositides and generates inositol 1,4,5-trisphosphate and diacyl-glycerol^[4]. This results in calcium mobilization and activation of protein kinase C (PKC)^[3]. Calcium is a neurotransmitter, and relatively low concentrations of calcium can cause a large response across the neuronal synapse^[3]. In addition to calcium stimulating an excitory response in nerve cells, PKC can be activated for regulatory purposes by the influx of calcium. A serine on PKC can become phosphorylated, which leaves PKC unable to bind to G beta-gamma (Gßγ) protein complex^[3]. Unbound Gßγ protein can then inhibit voltage-sensitive calcium channels to reduce calcium influx and provide feedback inhibition to the glutamate signaling pathway. ^[3].

Extracellular Domain

The extracellular domain of mGlu5 contains several key extracellular loops that will help modulate ligand binding. are shown here with extracellular loops (ECLs) 1, 2, and 3 highlighted in purple. Additionally in the ECL domain, a is attached to both Helix 3 and the amino acid chain between Helix 5 and the N-terminus. Helix 3 and Helix 5 are colored in teal and red respectively.The N-terminus is represented in blue. The disulfide bond is highlighted in yellow, and it is conserved in all classes of mGlu5 TMD^[2]. The disulfide bond is critical in maintaining the position of ECL 2 ^[1]. ECLs and the helices are also factors that dictate how mavoglurant fits in the binding pocket ^[1]. The position of these ECLs can change the effective size of the binding pocket through loop positioning^[1].

Binding Pocket

Figure 2. Mavoglurant in its binding pocket of the 7TM region of mGLu5 Class C receptor. The binding pocket's surface is clipped in black with the substrate, mavoglurant, in red. The rest of the protein is colored in green. The binding pocket is present in the 7 Helix Transmembrane Domain that would be present in the phospholipid bilayer as an integral protein. The presence of mavoglurant inhibits the function of the metabotropic glutamate receptor.

The binding pocket, not to be confused with the glutamate binding site, represents a useful source of regulatory control. The binding pocket is only accessible by a relatively narrow (7 Å) ^[1]. This small entrance severely restricts the access of both positive and negative allosteric regulators. One such regulator, , is a relatively small molecule that can travel through the entrance and fit between the helices of the mGlu5 receptor^[1]. Mavoglurant acts as a negative allosteric modulator of the mGlu5 receptor^[1]. Mavoglurant is a relatively hydrophobic molecule, which compliments the largely hydrophobic nature of the interior of mGlu5^[1]. The carbamate tail and hydroxyl group of mavoglurant will also interact with several key amino acids in the binding site of mGlu5^[1].

Important Amino Acids^[1]:

747 forms a hydrogen bond network with the main chain carbonyl of Glycine 652 and the carbamate portion of mavoglurant.
Bicyclic ring of mavoglurant surrounded by . Hydrophobic regions of the mGlu5 receptor are shown in purple and the rest of the helices are shown in grey. Mavoglurant is represented by spheres.
2 Catalytic residues H-bond to the hydroxyl oxygen of our ligand.
A inside of the binding pocket helps stabilize the inactive state.

Upon mavoglurant’s diffusion into the mGlu5 receptor binding pocket, the ordered water cage found in the center of mGlu5 is displaced^[1]. Favorable interactions between the hydrophobic regions of the binding pocket and the bicyclic ring and the 3-methylphenyl ring of mavoglurant help increase the strength and energetic favorability of mavoglurant binding via the hydrophobic effect. In the binding pocket mavoglurant can hydrogen bond with Asp 747 and that hydrogen bond will be further stabilized by an extended hydrogen bond network with Gly 652^[1]. The hydroxyl group of mavoglurant will also form another hydrogen bond with Ser 805 and Ser 809^[1]. Multiple hydrogen bonds make the binding of mavoglurant to the mGlu5 receptor favorable and specific^[2].

Once bound to mavoglurant, transmembrane helix 7 undergoes a conformational change^[1]. The shifting of TM7 will lead to a more global conformational change, which inactivates the receptor by moving intracellular loops inward ^[1]. This will leave the G-protien unable to bind with mGlu5^[2]. Variation can be seen in positioning of alpha helices 5 and 7 across receptor class. Class C receptors have less space for mavoglurant to enter as compared to Class A and F receptors^[2]. Increased specificity and stronger binding affinity could be a result of the more narrow structure of mGlu5.

Ionic Locks

Another important structural feature is the series of on the intracellular side of the protein. Interactions between these five amino acids will form a salt bridge, which will stabilize the inactive conformation^[1]. The primary ionic lock forms between Glu770, Lys665, and Ser613^[1]. A secondary ionic lock occurs between Ser614 and Arg668^[1]. The purpose of these ionic locks is analogous to the ionic interactions that stabilize the T state in Hemoglobin. In the case of the TMD of mGlu5, the ionic lock is formed when the NAM mavoglurant is bound. These stabilize the inactive state, where the intracellular loops are stabilized facing inwards^[2]. This conformational change will effectively block the crevice that is involved in binding the G-protein^[2]. Models have suggested that, even in a glutamate bound state, the mavoglurant bound receptor would be dimerized but incapable of signaling^[2]. This signaling incapable mGlu5 dimer will help maintain the readiness of the pathway, while still decreasing signal response.

Disease

Fragile X

Fragile X syndrome is the most common genetic cause of mental disability, and is a member of the Autism spectrum disorder family^[5]. The severity of intellectual disability can vary from patient to patient, but symptoms commonly stem from a misregulation of the mGlu1 and MGlu5 pathways^[5]. Misregulation of these pathways leads to over potentiation in neural cells. and other allosteric regulators like fenobam have shown promise in treating Fragile X. One positive characteristic of ligands that target the TMD of mGlu5 is they tend to be more specific, thus interacting less with nonspecific brain proteins^[6]. Mavoglurant down regulates glutamate signaling in an attempt to decrease potentiation. Unfortunately, recent Phase 2 clinical trials have proven mavoglurant ineffective ^[5]. However, modulators of mGlu5 TMD are still being investigated as treatment for Parkinson's, Alzheimer's disease, and various addictions^[3].

References

^[1] ^[2] ^[3] ^[6] ^[5] ^[4]

↑ ^1.00 ^1.01 ^1.02 ^1.03 ^1.04 ^1.05 ^1.06 ^1.07 ^1.08 ^1.09 ^1.10 ^1.11 ^1.12 ^1.13 ^1.14 ^1.15 ^1.16 ^1.17 ^1.18 ^1.19 ^1.20 ^1.21 ^1.22 ^1.23 ^1.24 ^1.25 ^1.26 ^1.27 ^1.28 Dore AS, Okrasa K, Patel JC, Serrano-Vega M, Bennett K, Cooke RM, Errey JC, Jazayeri A, Khan S, Tehan B, Weir M, Wiggin GR, Marshall FH. Structure of class C GPCR metabotropic glutamate receptor 5 transmembrane domain. Nature. 2014 Jul 31;511(7511):557-62. doi: 10.1038/nature13396. Epub 2014 Jul 6. PMID:25042998 doi:http://dx.doi.org/10.1038/nature13396
↑ ^2.00 ^2.01 ^2.02 ^2.03 ^2.04 ^2.05 ^2.06 ^2.07 ^2.08 ^2.09 ^2.10 ^2.11 ^2.12 ^2.13 ^2.14 ^2.15 Wu H, Wang C, Gregory KJ, Han GW, Cho HP, Xia Y, Niswender CM, Katritch V, Meiler J, Cherezov V, Conn PJ, Stevens RC. Structure of a class C GPCR metabotropic glutamate receptor 1 bound to an allosteric modulator. Science. 2014 Apr 4;344(6179):58-64. doi: 10.1126/science.1249489. Epub 2014 Mar , 6. PMID:24603153 doi:http://dx.doi.org/10.1126/science.1249489
↑ ^3.0 ^3.1 ^3.2 ^3.3 ^3.4 ^3.5 ^3.6 ^3.7 ^3.8 Niswender CM, Conn PJ. Metabotropic glutamate receptors: physiology, pharmacology, and disease. Annu Rev Pharmacol Toxicol. 2010;50:295-322. doi:, 10.1146/annurev.pharmtox.011008.145533. PMID:20055706 doi:http://dx.doi.org/10.1146/annurev.pharmtox.011008.145533
↑ ^4.0 ^4.1 Woodcock EA, Kistler PM, Ju YK. Phosphoinositide signalling and cardiac arrhythmias. Cardiovasc Res. 2009 May 1;82(2):286-95. doi: 10.1093/cvr/cvn283. Epub 2008 Oct, 20. PMID:18940816 doi:http://dx.doi.org/10.1093/cvr/cvn283
↑ ^5.0 ^5.1 ^5.2 ^5.3 Bailey DB Jr, Berry-Kravis E, Wheeler A, Raspa M, Merrien F, Ricart J, Koumaras B, Rosenkranz G, Tomlinson M, von Raison F, Apostol G. Mavoglurant in adolescents with fragile X syndrome: analysis of Clinical Global Impression-Improvement source data from a double-blind therapeutic study followed by an open-label, long-term extension study. J Neurodev Disord. 2016;8:1. doi: 10.1186/s11689-015-9134-5. Epub 2015 Dec 15. PMID:26855682 doi:http://dx.doi.org/10.1186/s11689-015-9134-5
↑ ^6.0 ^6.1 Feng Z, Ma S, Hu G, Xie XQ. Allosteric Binding Site and Activation Mechanism of Class C G-Protein Coupled Receptors: Metabotropic Glutamate Receptor Family. AAPS J. 2015 May;17(3):737-53. doi: 10.1208/s12248-015-9742-8. Epub 2015 Mar 12. PMID:25762450 doi:http://dx.doi.org/10.1208/s12248-015-9742-8

External Resources

Novartis Fragile X trials

Mavoglurant as a treatment for Parkinson's disease

Pharmacology of mGlu5 receptors

Glutamate signaling pathway and receptors general overview

Gq Alpha subunit

Retrieved from "http://52.214.119.220/wiki/index.php/Sandbox_Reserved_1160"

@@ Line 4: / Line 4: @@
 The functionality of the mGlu5 receptor is determined by conformational changes throughout multiple domains. mGlu5 will bind glutamate through its extracellular Venus flytrap domain and the signal will be transduced across the membrane to a heterotrimeric G protein, which will ultimately lead to calcium release and the activation of [[PKC]]<ref name="Wu" />. The signal is relayed through a Gq/11 pathway<ref name="Dore" />. Activated PKC will elicit a excitory post-synaptic response and modulate long term potentiation in the CNS<ref name="Wu" />.
-Human mGlu5 is found throughout the central nervous system. Areas containing high concentrations of mGlu5 are often involved in emotional processing and higher cognition<ref name="Niswender" />. The localization of mGlu5 in the CNS and the presence of multiple domains makes mGlu5 an excellent target for treating neurological conditions including: schizophrenia,[http://www.fragilex.org/fragile-x/fragile-x-syndrome/ Fragile X], [http://www.en.wikipedia.org/wiki/Depression_(mood)depression], [http://www.en.wikipedia.org/wiki/Anxiety_disorder anxiety], and [http://www.en.wikipedia.org/wiki/Alzheimer's_disease Alzheimer's disease]<ref name="Wu" />.
+Human mGlu5 is found throughout the central nervous system. Areas containing high concentrations of mGlu5 are often involved in emotional processing and higher cognition<ref name="Niswender" />. The localization of mGlu5 in the CNS and the presence of multiple domains makes mGlu5 an excellent target for treating neurological conditions including: schizophrenia, [http://www.fragilex.org/fragile-x/fragile-x-syndrome/ Fragile X], [http://www.en.wikipedia.org/wiki/Depression_(mood)depression], [http://www.en.wikipedia.org/wiki/Anxiety_disorder anxiety], and [http://www.en.wikipedia.org/wiki/Alzheimer's_disease Alzheimer's disease]<ref name="Wu" />.
 == Discovery ==
-The mGlu family of receptors was the first of the Class C [[GPCR]] to be extensively studied<ref name="Wu" />. The first regions of the protein crystallized and studied were the Venus Fly Trap domain and the cysteine -rich domain on the extracellular region of the receptor<ref name="Dore" />.The Venus Flytrap domain is a large extracellular domain that will selectively bind to glutamate<ref name="Wu" />. The CRD is a somewhat smaller domain composed of many ß sheets and cysteine residues <ref name="Wu" />. The CRD acts as a signal mediator between the Venus Flytrap domain and the TMD of mGlu5, by linking to each domain with disulfide bonds<ref name="Wu" />. The hydrophobic nature and flexibility of the mGlu5 TMD made it difficult to crystallize.
+The mGlu family of receptors was the first of the Class C [[GPCR]] to be extensively studied<ref name="Wu" />. The first regions of the protein crystallized and studied were the Venus Fly Trap domain and the cysteine -rich domain (CRD) on the extracellular region of the receptor<ref name="Dore" />.The Venus Fly Trap domain is a large extracellular domain that will selectively bind to glutamate<ref name="Wu" />. The CRD is a somewhat smaller domain composed of many ß sheets and cysteine residues <ref name="Wu" />. The CRD acts as a signal mediator between the Venus Flytrap domain and the transmembrane domain (TMD) of mGlu5, by linking to each domain with disulfide bonds<ref name="Wu" />. The hydrophobic nature and flexibility of the mGlu5 TMD made it difficult to crystallize.
-Recently, the human metabotropic glutamate receptor 5 transmembrane domain (TMD) was crystallized and a structure elucidated<ref name="Dore" />. Several modifications were made to the TMD for successful crystallization. The protein was thermostabilized and flexible domains were removed<ref name="Dore" />. In total residues 2-568 and 837-1153 were excised from the structure. These flexible domains allow mGlu5 to bind to its GPCR <ref name="Dore" />. The structure of the alpha helices are shown in <font color='darkgreen'>'''green'''</font>, and the negative allosteric modulator mavoglurant shown in <span style="color:yellow;background-color:black;font-weight:bold;">yellow</span>. Also in <font color='orange'><b>orange</b></font>, a T4 -<scene name='72/721531/Protien_lys/3'>Lysozyme</scene> was inserted into intracellular loop 2 to add stability<ref name="Dore" />.
+Recently, the human metabotropic glutamate receptor 5 transmembrane domain was crystallized and a structure elucidated<ref name="Dore" />. Several modifications were made to the TMD for successful crystallization. The protein was thermostabilized and flexible domains were removed<ref name="Dore" />. In total residues 2-568 and 837-1153 were excised from the structure. These flexible domains allow mGlu5 to bind to its GPCR <ref name="Dore" />. The structure of the alpha helices are shown in <font color='darkgreen'>'''green'''</font>, and the negative allosteric modulator mavoglurant shown in <span style="color:yellow;background-color:black;font-weight:bold;">yellow</span>. Also in <font color='orange'><b>orange</b></font>, a T4 -<scene name='72/721531/Protien_lys/3'>Lysozyme</scene> was inserted into intracellular loop 2 to add stability<ref name="Dore" />.
 == Structure==
 [[Image:STR.png|200 px|left|thumb|'''Figure 1''': Overall Structure of the mGlu5 TMD. The polar heads on the Oleic acids orient the image with the top oriented extracellularly, the middle portion inserted into the membrane, and the lower portion oriented intracellularly. The white exterior represents the surface of the protien, and the multicolored lines interior to the surface represent the backbones 7 transmembrane alpha helices. ]]
 === Overview ===
-The mGlu5 TMD contains 7 <scene name='72/721531/Protien_7_helices/4'> alpha helices</scene> that spans the membrane. The protein was crystallized with <scene name='72/721531/Protien_clean_sce/6'>Oleic acid</scene> shown in <font color='red'>'''red'''</font>. On the top portion of the protein several critical extracellular loops connect the TMD to the CRD. The binding pocket can be found deep in the interior of the protein, and is mainly comprised of hydrophobic amino acids with more polar amino acids found in the upper and lower portions of the binding site<ref name="Dore" />. Inserted into the biding pocket is the negative allosteric modulator [http://www.en.wikipedia.org/wiki/Mavoglurant mavoglurant]. The TMD is in an inactive conformation, since mavoglurant is bound<ref name="Dore" />. Also, the deletion of the flexible domains leaves the mGlu5 receptor unable to bind to its [[GPCR]]<ref name="Dore" />. The inactive state is maintained by multiple ionic locks whose positions determine the active versus inactive conformation.
+The mGlu5 TMD contains 7 <scene name='72/721531/Protien_7_helices/4'> alpha helices</scene> that spans the membrane. The protein was crystallized with <scene name='72/721531/Protien_clean_sce/6'>Oleic acid</scene>  shown in <font color='red'>'''red'''</font>. On the top portion of the protein several critical extracellular loops connect the TMD to the CRD. The binding pocket can be found deep in the interior of the protein, and is mainly comprised of hydrophobic amino acids with more polar amino acids found in the upper and lower portions of the binding site<ref name="Dore" />. Inserted into the biding pocket is the negative allosteric modulator [http://www.en.wikipedia.org/wiki/Mavoglurant mavoglurant]. The TMD is in an inactive conformation, since mavoglurant is bound<ref name="Dore" />. Also, the deletion of the flexible domains leaves the mGlu5 receptor unable to bind to its [[GPCR]]<ref name="Dore" />. The inactive state is maintained by multiple ionic locks whose positions determine the active versus inactive conformation.
-When mGlu5 is in the active conformation, signaling begins with glutamate binding to the Venus flytrap domain. The signal is transduced across the cysteine-rich domain to the TMD<ref name="Niswender" />. Next, the dimerization of the TMD occurs. This activates the Gq/11 pathway, which activates phospholipase Cβ<ref name="Niswender" />. The active [http://www.proteopedia.org/wiki/index.php/2zkm phospholipase Cβ] hydrolyzes phosphotinositides and generates [https://pubchem.ncbi.nlm.nih.gov/compound/439456#section=Top inositol 1,4,5-trisphosphate] and [http://www.sivabio.50webs.com/ip3.htm diacyl-glycerol]<ref name="Woodcock" />. This results in calcium mobilization and activation of protein kinase C ([[PKC]])<ref name="Niswender" />. Calcium is a neurotransmitter, and relatively low concentrations of calcium can cause a large response across the neuronal synapse<ref name="Niswender" />. In addition to calcium stimulating an exitory response in nerve cells, PKC can be activated for regulatory purposes by the influx of calcium. A serine on PKC can become phosphorylated, which leaves PKC unable to bind to G beta-gamma (Gßγ) protein complex<ref name="Niswender" />. Unbound Gßγ protein can then inhibit voltage-sensitive calcium channels to reduce calcium influx and provide feedback inhibition to the glutamate signaling pathway. <ref name="Niswender" />.
+When mGlu5 is in the active conformation, signaling begins with glutamate binding to the Venus flytrap domain. The signal is transduced across the cysteine-rich domain to the TMD<ref name="Niswender" />. Next, the dimerization of the TMD occurs. This activates the Gq/11 pathway, which activates phospholipase Cβ<ref name="Niswender" />. The active  [http://www.proteopedia.org/wiki/index.php/2zkm phospholipase Cβ] hydrolyzes phosphotinositides and generates [https://pubchem.ncbi.nlm.nih.gov/compound/439456#section=Top inositol 1,4,5-trisphosphate] and [http://www.sivabio.50webs.com/ip3.htm diacyl-glycerol]<ref name="Woodcock" />. This results in calcium mobilization and activation of protein kinase C ([[PKC]])<ref name="Niswender" />. Calcium is a neurotransmitter, and relatively low concentrations of calcium can cause a large response across the neuronal synapse<ref name="Niswender" />. In addition to calcium stimulating an excitory response in nerve cells, PKC can be activated for regulatory purposes by the influx of calcium. A serine on PKC can become phosphorylated, which leaves PKC unable to bind to G beta-gamma (Gßγ) protein complex<ref name="Niswender" />. Unbound Gßγ protein can then inhibit voltage-sensitive calcium channels to reduce calcium influx and provide feedback inhibition to the glutamate signaling pathway. <ref name="Niswender" />.
 === Extracellular Domain ===
 The extracellular domain of mGlu5 contains several key extracellular loops that will help modulate ligand binding. <scene name='72/721532/Ecl_trail_7/2'>Extracellular Loops</scene> are shown here with extracellular loops (ECLs) <font color='purple'><b>1</b></font>, <font color='purple'><b>2</b></font>, and <font color='purple'><b>3</b></font> highlighted in <font color='purple'><b>purple</b></font>. Additionally in the ECL domain, a <scene name='72/721531/Ecl_trail_1/5'>disulfide bond</scene> is attached to both Helix 3 and the amino acid chain between Helix 5 and the N-terminus. <font color='teal'><b>Helix 3</b></font> and <font color='red'><b>Helix 5</b></font> are colored in <font color='teal'><b>teal</b></font> and <font color='red'><b>red</b></font> respectively.The <font color='blue'><b>N-terminus</b></font> is represented in <font color='blue'><b>blue</b></font>. The <span style="color:yellow;background-color:black;font-weight:bold;">disulfide bond</span> is highlighted in <span style="color:yellow;background-color:black;font-weight:bold;">yellow</span>, and it is conserved in all classes of mGlu5 TMD<ref name="Wu" />. The disulfide bond is critical in maintaining the position of ECL 2 <ref name="Dore" />. ECLs and the helices are also factors that dictate how mavoglurant fits in the binding pocket <ref name="Dore" />. The position of these ECLs can change the effective size of the binding pocket through loop positioning<ref name="Dore" />.
@@ Line 37: / Line 37: @@
 == Disease ==
 === Fragile X ===
-Fragile X syndrome is the most common genetic cause of mental disability, and is a member of the [http://www.nimh.nih.gov/health/topics/autism-spectrum-disorders-asd/index.shtml Autism spectrum disorder] family<ref name="Bailey" />. The severity of intellectual disability can vary from patient to patient, but symptoms commonly stem from a misregulation of the mGlu1 and MGlu5 pathways<ref name="Bailey" />. Misregulation of these pathways leads to over potentiation in neural cells. <scene name='72/721531/Mavo/2'>Mavoglurant</scene> and other allosteric regulators like [http://www.en.wikipedia.org/wiki/Fenobam fenobam] have shown promise in treating Fragile X. One positive characteristic of ligands that target the TMD of mGlu5 is they tend to be more specific, thus interacting less with nonspecific brain proteins<ref name="Feng" />. Mavoglurant down regulates glutamate signaling in an attempt to decrease potentiation. Unfortunately, recent Phase 2 clinical trials have proven mavoglurant ineffective <ref name="Bailey" />.  However, modulators of mGlu5 TMD are still being investigated as treatment for to Parkinson's, Alzheimer's disease, and various addictions<ref name="Niswender" />.
+Fragile X syndrome is the most common genetic cause of mental disability, and is a member of the [http://www.nimh.nih.gov/health/topics/autism-spectrum-disorders-asd/index.shtml Autism spectrum disorder] family<ref name="Bailey" />. The severity of intellectual disability can vary from patient to patient, but symptoms commonly stem from a misregulation of the mGlu1 and MGlu5 pathways<ref name="Bailey" />. Misregulation of these pathways leads to over potentiation in neural cells. <scene name='72/721531/Mavo/2'>Mavoglurant</scene> and other allosteric regulators like [http://www.en.wikipedia.org/wiki/Fenobam fenobam] have shown promise in treating Fragile X. One positive characteristic of ligands that target the TMD of mGlu5 is they tend to be more specific, thus interacting less with nonspecific brain proteins<ref name="Feng" />. Mavoglurant down regulates glutamate signaling in an attempt to decrease potentiation. Unfortunately, recent Phase 2 clinical trials have proven mavoglurant ineffective <ref name="Bailey" />.  However, modulators of mGlu5 TMD are still being investigated as treatment for Parkinson's, Alzheimer's disease, and various addictions<ref name="Niswender" />.