<StructureSection load='4l6r' size='350' side='right' caption='Structure of the Class B Human Glucagon G Protein Coupled Receptor-[http://www.rcsb.org/pdb/home/home.do PDB] [http://www.rcsb.org/pdb/explore/explore.do?structureId=4l6r 4L6R]' scene='72/721536/Class_b_gpcrs/3'>
==Human Glucagon Class B G Protein-Coupled Receptors (GPCRs)==

===Introduction===
[[Image:Screen_Shot_2016-03-29_at_12.45.02_PM.png|(|):|200 px|right|thumb|<font size="1.0"><div style="text-align: center;">'''Figure 1: Glucagon''', shown in green, bound to its receptor in the class B family </div></font>]]
'''Human glucagon class B G protein-coupled receptors (GPCRs'''), also known as [https://en.wikipedia.org/wiki/Secretin_receptor_family secretin-like receptors], are a subfamily GPCRs and very similar in structure to the more well known class A ([https://en.wikipedia.org/wiki/Rhodopsin-like_receptors rhodopsin-like]) glucagon receptor family. <ref name="Intro">PMID: 24359917</ref> Located in the [https://en.wikipedia.org/wiki/Liver liver], class B glucagon receptors (GCGRs) are activated by the binding of the hormonal peptide [https://en.wikipedia.org/wiki/Glucagon glucagon] (Figure 1). Glucagon binding leads to the release of [https://en.wikipedia.org/wiki/Glucose glucose] into the [https://en.wikipedia.org/wiki/Circulatory_system bloodstream] and plays an essential role in [https://en.wikipedia.org/wiki/Blood_sugar_regulation glucose homeostasis]. Class B GCGRs are composed of a [https://en.wikipedia.org/wiki/Liver seven transmembrane domain] (7TM) and [https://en.wikipedia.org/wiki/Liver extracellular domain] (ECD) that are required for glucagon binding.

=Structures of Class A vs. Class B GPCRs=
In comparison, class A vs. class B glucagon receptors share less than fifteen percent sequence homology, but both share this 7TM domain. <ref name="Intro">PMID: 24359917</ref> Understanding for class A family of GCGRs for the structure-function [https://en.wikibooks.org/wiki/Structural_Biochemistry/Enzyme_Catalytic_Mechanism mechanism] has made great progress over the past few years, but understanding of class B has fallen behind but has recently started catching up. <ref name="Tips">PMID: 23863937</ref> Comparison of the <scene name='72/721536/7tm/2'>Class B GCGR 7TM</scene> was compared to that of a <scene name='72/721536/Class_a/1'>class A 7TM</scene>, and it was found that the orientation and positioning of the [https://en.wikipedia.org/wiki/Alpha_helix alpha helices] are conserved through both classes. But, structural alignments of the two revealed multiple gaps in the transmembrane region signifying a variety of structural deviations in transmembrane helices. <ref name="Tips">PMID: 23863937</ref. The N-terminal end of helix one in class B GCGR, located in the 7TM, is longer than any known class A GPCR structure and stretches three supplementary helical turns above the extracellular (EC) membrane boundary. This region is referred to as the <scene name='72/721535/Opening_orientation/2'>stalk</scene> and is involved in glucagon binding and helps in defining the orientation of the ECD with respect to the 7TM domain <ref name="Tips">PMID: 23863937</ref>. Also specific to class B GPCRs, a [https://en.wikipedia.org/wiki/Glycine glycine] (Gly) residue at position 393 induces a <scene name='72/721535/Helical_bend/2'>bend in helix VII</scene>; this bend is stabilized by the [http://chemwiki.ucdavis.edu/Core/Physical_Chemistry/Physical_Properties_of_Matter/Atomic_and_Molecular_Properties/Intermolecular_Forces/Hydrophobic_Interactions hydrophobic interaction] between the <scene name='72/721535/Gly_393_phe_184/1'>glycine 393 and phenylalanine 184</scene>. One of the most distinguishable characteristics of the class B 7TM is the <scene name='72/721536/Helix_eight_tilt/2'>helix VIII tilt</scene> of 25 degrees compared to that of class A, which has no tilt. This results from a [https://en.wikipedia.org/wiki/Phenylalanine glutamate] (Glu) 406 in helix VIII that is fully conserved in secretin-like receptors and forms two interhelical [https://en.wikipedia.org/wiki/Salt_bridge_(protein_and_supramolecular) salt bridges] with [https://simple.wikipedia.org/wiki/Conserved_sequence conserved residues] [https://en.wikipedia.org/wiki/Arginine arginine] (Arg) 173 and arginine 346 <ref name="Tips">PMID: 23863937</ref>. Despite these differences, a vital region that is conserved in both class B and class A receptors is the [https://en.wikipedia.org/wiki/Disulfide disulphide bond] between [https://en.wikipedia.org/wiki/Cysteine cysteine] <scene name='72/721535/Disulfide_bond_notspin/1'>(Cys) 294 and cysteine 224</scene> in extracellular loop two (ECL2). This bond stabilizes the receptors entire 7TM fold. Lastly, the locations of the extracellular tips for class B glucagon receptors allow for a much wider and deeper binding cavity in the [https://en.wikipedia.org/wiki/Ligand_(biochemistry) ligand-binding pocket], which is much more immense than any of the class A GCGRs <ref name="Tips">PMID: 23863937</ref>. These wide ranges specifically occur between two sets of alpha helices, (Figure 2).

[[Image:Colored_Helices_for_Proteopedia.png|(|):|350 px|center|thumb|'''Figure 2: Extracellular tips of the 7TM helices.''' Helices two and six are shown in green, while helices three and seven are shown in red]]

==How These Structures Lead to Function==
Structurally, the [https://en.wikipedia.org/wiki/N-terminus N-terminal] <scene name='72/721535/Ecd/2'>extracellular domain (ECD)</scene> and the 7TM comprise the signature seven helical structure that is involved in [https://en.wikibooks.org/wiki/Principles_of_Biochemistry/Signaling_inside_the_Cell signaling] via [https://en.wikibooks.org/wiki/Structural_Biochemistry/Energy_coupling_in_chemical_reactions coupling] to [https://en.wikipedia.org/wiki/Heterotrimeric_G_protein heterotrimeric G proteins] that activate [https://en.wikipedia.org/wiki/Adenylyl_cyclase adenylate cyclase] to increase the levels of intracellular [https://en.wikipedia.org/wiki/Cyclic_adenosine_monophosphate cyclic AMP]. Additionally, this coupling increases [https://en.wikipedia.org/wiki/Inositol_phosphate inositol phosphate] and intracellular [https://en.wikipedia.org/wiki/Calcium calcium] levels <ref name="Tips">PMID: 23863937</ref>. The wider and deeper ligand-binding pocket of class B GPCRs allows for a vast array of [https://en.wikipedia.org/wiki/Receptor_(biochemistry) receptors] to be bound that allow for numerous functions activated by peptide receptors <ref name="Ligands">PMID: 21542831</ref>. The conformation and orientation of the 7TM and the ECD regions dictate the functionality of the protein, which has an open and closed [https://en.wikipedia.org/wiki/Conformation conformation] of the GCGR. When glucagon binds to GCGR, the open conformation of GCGR is stabilized. There is no clear [https://en.wikipedia.org/wiki/Active_site binding site] location of the hormone peptide ligand, but they do know the N-terminus of glucagon binds deep into the <scene name='72/721535/Binding_pocket_orange/1'>binding pocket</scene>. The [https://en.wikipedia.org/wiki/Amino_acid amino acids] at the N-terminus have the ability to form [https://en.wikipedia.org/wiki/Hydrogen_bond hydrogen bonds] and [https://en.wikipedia.org/wiki/Ionic_bonding ionic interactions] involved, which can be seen in the [https://en.wikipedia.org/wiki/Peptide_sequence amino acid sequence] of glucagon (Figure 3). <ref name="Sequence">PMID: 11946536</ref>.

[[Image:Aminoacidsequenceglucagon.png|(|):|425 px|center|thumb|'''Figure 3: Amino Acid Sequence of Glucagon''']]

Many of the [https://en.wikipedia.org/wiki/Residue_(chemistry) residues] that are in direct contact with the glucagon molecule are [https://en.wikipedia.org/wiki/Ion charged] or are [https://en.wikipedia.org/wiki/Chemical_polarity polar].
[[Image:Screen Shot 2016-03-29 at 3.24.43 PM.png|(|):|400 px|right|thumb|'''Figure 4: Salt Bridge'''. The salt bridge is located on the intracellular side at the bottom of the protein, in relation to the orientation it holds within the cell membrane. It is made between residues Glu 406, Arg 173, and Arg 346, as labeled in the figure.]]
There are also many smaller residues on glucagon that support the bulky residues on the GCGR. These residues are located within the binding pocket of the 7TM <ref name="Ligands">PMID: 21542831</ref>. There are specific amino acid interactions that hold the helices of the 7TM in the closed conformation that maximizes [http://www.chemicool.com/definition/affinity.html affinity]. This includes the [https://en.wikipedia.org/wiki/Disulfide disulfide bond] between Cys 294 and Cys 224 that was mentioned earlier that serves to hold the ECL1 and ECL2 in the proper orientation. Additionally, the [https://en.wikipedia.org/wiki/Salt_bridge_%28protein_and_supramolecular%29 salt bridges] between Glu 406, Arg 173, and Arg 346, also mentioned earlier, hold the conformation together for higher affinity. Finally, alpha helical structure of the stalk is imperative to the affinity and binding of the glucagon <ref name="Tips">PMID: 23863937</ref>.

[[Image:Screen_Shot_2016-03-22_at_5.28.03_PM.png|(|):|425 px|center|thumb|'''Figure 5: Binding Pocket Residues:''' side chains of carbon chains are shown in green and side chains containing oxygen are shown in red]]

=Glucagon Signaling Pathway=
For glucagon to initiate the breakdown of [https://en.wikipedia.org/wiki/Glycogen glycogen] into glucose, it follows a specific [https://en.wikipedia.org/wiki/Glucagon signaling mechanism] (Figure 6). Glucagon binds to a G protein-coupled receptor located on the [https://en.wikipedia.org/wiki/Cell_membrane plasma membrane], and this binding induces a [https://en.wikipedia.org/wiki/Conformational_change conformational change] that activates G proteins which exchanges a [https://en.wikipedia.org/wiki/Guanosine_diphosphate guanosine diphosphate (GDP]) for guanosine [https://en.wikipedia.org/wiki/Guanosine_triphosphate triphosphate (GTP)] that is bound to the [https://en.wikipedia.org/wiki/G_alpha_subunit alpha subunit]. Now with the GTP in place, the activated alpha subunit leaves the [https://en.wikipedia.org/wiki/Heterotrimeric_G_protein heterotrimeric G protein] and activatesthe next [https://en.wikipedia.org/wiki/Enzyme enzyme] in the cascade, [https://en.wikipedia.org/wiki/Adenylyl_cyclase adenylate cyclase]. Once adenylate cyclase is activated, it catalyzes the conversion of [https://en.wikipedia.org/wiki/Adenosine_triphosphate adenosine triphosphate (ATP)] into cyclic [https://en.wikipedia.org/wiki/Cyclic_adenosine_monophosphate adenosine monophosphate (cAMP)]. This in turn activates [https://en.wikipedia.org/wiki/Protein_kinase_A protein kinase A (PKA)] that then activates [https://en.wikipedia.org/wiki/Phosphorylase_kinase phosphorylase kinase]. Finally, the phosphorylase kinase [https://en.wikipedia.org/wiki/Phosphorylation phosphorylates] [https://en.wikipedia.org/wiki/Glycogen_phosphorylase glycogen phosphorylase b] that is then turned into its activated form, glycogen phosphorylase a. This phosphorylase A enzyme is responsible for catalyzing the release of [https://en.wikipedia.org/wiki/Glucose_1-phosphate glucose-1-phosphate] into the bloodstream from glycogen [https://en.wikipedia.org/wiki/Polymer polymers].

[[Image:Glucagon_Pathway.png|(|):|400 px|center|thumb|'''Figure 6: [https://en.wikipedia.org/wiki/Glucagon Glucagon Signaling Pathway]''']]

=Clinical Relevancy=
Of the fifteen human class B GPCRs, eight have been identified as potential [https://en.wikipedia.org/wiki/Biological_target drug target]<ref name="Drug">PMID: 24628305</ref>. [http://www.wisegeek.com/what-are-therapeutic-agents.htm Therapeutic agents] have been created from the peptides themselves within this protein, but overall [https://en.wikipedia.org/wiki/Pharmaceutical_industry pharmaceutical companies] have had difficulty creating agents that act on family B GPCRS. There is an outward appearance and inherent flexibility in the class B GCGR 7TM because of conserved hydrogen bonds that flank a glycine residue, and this structure along with the ECD and its role of interactions on the extracellular side of receptors may provide evidence to how class B receptors adjust its conformational spectra for various receptors. Researchers hope to show how these conformations can be utilized in potential treatments of a wide array [https://en.wikipedia.org/wiki/List_of_mental_disorders disorders].
==Potential Inhibitors==
Research for class B GCGR [https://en.wikipedia.org/wiki/Enzyme_inhibitor inhibitors] is primarily looking into [https://en.wikipedia.org/wiki/Allosteric_regulation allosteric inhibitors] having the ability to target specific receptors in order to treat problems like [https://en.wikipedia.org/wiki/Stress-related_disorders stress disorders], managing [http://www.webmd.com/diabetes/guide/diabetes-hyperglycemia hyperglycemia], and also alternative mechanisms for treating [https://en.wikipedia.org/wiki/Migraine migraines] <ref name="Inhibitors">PMID: 24189067</ref>. Known inhibitors include [https://en.wikipedia.org/wiki/Monoclonal_antibody monoclonal antibodies] which inhibit glucagon receptors through an allosteric mechanism. The monoclonal antibodies bind to two different sites, the ECD opposite of the binding region and then the helical portion of the ECD as well. <ref name="Last">PMID: 19305799</ref>.
===Research===
Determining the structure of class B GCGRs is a reason for its lack of advanced knowledge in the field, but [https://en.wikipedia.org/wiki/X-ray_crystallography X-ray crystallography] and [https://en.wikipedia.org/wiki/Nuclear_magnetic_resonance NMR] have been the main processes performed and have had some success with it over the past couple years <ref name="Lastt">PMID: 26227798</ref>. X-ray crystallography displayed the [https://en.wikipedia.org/wiki/Crystal_structure crystal structure] of ECDs of class B GPCRs in complex with their [https://en.wikipedia.org/wiki/Ligand ligands] along with the crystal structure of the 7TM. In addition to this, NMR has allowed the ability to directly understand structures of soluble amino-terminal domains of numerous members of the secretin-like family that bind [https://en.wikipedia.org/wiki/Peptide_hormone peptide hormones]. Primary sequences analysis have led to the finding of seven segments of eighteen or more relatively hydrophobic residues that are believed to represent transmembrane helices that take part in creating an intramembranous helical bundle <ref name="Lastt">PMID: 26227798</ref>. Also, [https://en.wikipedia.org/wiki/Mutagenesis mutagenesis] has been used to determine which residues were necessary in maximizing affinity for glucagon. Finally, the orientation and mechanism of the peptide interactions within these structures are studied using peptide structure-activity relationships (SAR), receptor and ligand fragments, chimeric receptors, site-directed mutagenesis, photochemical cross-linking, and molecular modeling <ref name="Lastt">PMID: 26227798</ref>.
</StructureSection>

== References ==
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