Andrew Helmerich Sandbox 1: Difference between revisions
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= | =Amylin Receptor= | ||
<StructureSection load='7tyf' size='340' side='right' caption='AMYR Bound to Amylin' scene='10/1038828/Entire_protein_scene/7'> | <StructureSection load='7tyf' size='340' side='right' caption='AMYR Bound to Amylin' scene='10/1038828/Entire_protein_scene/7'> | ||
==Introduction== | ==Introduction== | ||
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Within the transmembrane domain (TMD) of the CTR, hydrophobic R groups span the phospholipid bilayer, anchoring the protein into the cell membrane upon amylin binding to the receptor. The interior of the transmembrane domain contains the hydrophilic residues necessary for ligand binding and transduction of the signal across the cell membrane. | Within the transmembrane domain (TMD) of the CTR, hydrophobic R groups span the phospholipid bilayer, anchoring the protein into the cell membrane upon amylin binding to the receptor. The interior of the transmembrane domain contains the hydrophilic residues necessary for ligand binding and transduction of the signal across the cell membrane. | ||
==== G-alpha Interactions with CTR TMD ==== | ==== G-alpha Interactions with CTR TMD ==== | ||
To transduce the signal across the cell membrane, the binding of amylin will induce a conformational change that allows for the CTR to make favorable interactions with the G alpha subunit. Two interactions shown <scene name='10/1038828/Ctr_g_alpha/15'>(1, </scene><scene name='10/1038828/Ctr_g_alpha/12'>2) </scene> activate the G-protein and propel downstream signaling. As with a typical glucagon GPCR pathway, the G-alpha subunit becomes activated upon guanine exchange factor [https://en.wikipedia.org/wiki/Guanine_nucleotide_exchange_factor (GEF)] echanging GDP for GTP. This G-alpha subunit transverses laterally in the membrane towards adenylyl cyclase, activating it and causing an increase in the second messenger cyclic adenosine monophosphate [https://en.wikipedia.org/wiki/Cyclic_adenosine_monophosphate (cAMP)]. This cAMP activates protein kinase A [https://en.wikipedia.org/wiki/Protein_kinase_A (PKA)], which can phosphorylate other proteins facilitating cellular response. | To transduce the signal across the cell membrane, the binding of amylin will induce a conformational change that allows for the CTR to make favorable interactions with the G alpha subunit. Two interactions shown <scene name='10/1038828/Ctr_g_alpha/15'>(1, </scene><scene name='10/1038828/Ctr_g_alpha/12'>2) </scene> activate the G-protein and propel downstream signaling. As with a typical glucagon GPCR pathway, the G-alpha subunit becomes activated upon guanine exchange factor [https://en.wikipedia.org/wiki/Guanine_nucleotide_exchange_factor (GEF)] echanging GDP for GTP. This G-alpha subunit transverses laterally in the membrane towards adenylyl cyclase, activating it and causing an increase in the second messenger cyclic adenosine monophosphate [https://en.wikipedia.org/wiki/Cyclic_adenosine_monophosphate (cAMP)]. This cAMP activates protein kinase A [https://en.wikipedia.org/wiki/Protein_kinase_A (PKA)], which can phosphorylate other proteins facilitating cellular response. | ||
====N-Terminus Disulfide==== | ====N-Terminus Disulfide==== | ||
The amylin peptide contains a <scene name='10/1038819/N_term_disulfide/ | The amylin peptide contains a <scene name='10/1038819/N_term_disulfide/4'>covalent disulfide linkage</scene> between residues C2 and C7. This disulfide provides stability and rigidity to the helical structure of the peptide, allowing for favorable binding to the extracellular domain (ECD). New interactions due to the disulfide include hydrogen bonds between E294 of the transmembrane domain with K1 of amylin, and both R362 and W361 of the transmembrane domain forming a hydrogen bond with N3 of amylin. | ||
====Amidated C-Terminus==== | ====Amidated C-Terminus==== | ||
The <scene name='10/1038819/Amidated_c_term/9'>C-Terminus</scene> of amylin contains an amide group, rather than a carboxylic acid group. This chemical modification allows for more extensive hydrogen bonding to nearby residues, due to the added hydrogen bond donor on the NH2 group. In turn, this allows for favorable hydrogen bonds between S129 of the transmembrane domain and the main chain of Y37 on amylin. This interaction causes a "kink" in the random coil of amylin, displacing Y37 into a hydrophobic pocket, allowing for favorable hydrophobic interactions with W79 of the transmembrane domain. This amidation is thought to be a post-translational modification. <ref name="Vekic">PMID: 36005584</ref> | The <scene name='10/1038819/Amidated_c_term/9'>C-Terminus</scene> of amylin contains an amide group, rather than a carboxylic acid group. This chemical modification allows for more extensive hydrogen bonding to nearby residues, due to the added hydrogen bond donor on the NH2 group. In turn, this allows for favorable hydrogen bonds between S129 of the transmembrane domain and the main chain of Y37 on amylin. This interaction causes a "kink" in the random coil of amylin, displacing Y37 into a hydrophobic pocket, allowing for favorable hydrophobic interactions with W79 of the transmembrane domain. This amidation is thought to be a post-translational modification. <ref name="Vekic">PMID: 36005584</ref> | ||
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It is hypothesized that amylin binds to the receptor via a two-domain model (Figure 2). The model suggests a series of steps for how amylin binds. First, the c-terminus of amylin binds to the n terminus of the extracellular domain of the receptor. This binding factors the alignment of amylin's n-terminus to the primary GPCR binding site. Once both the c-terminus and n-terminus of amylin are bound, the receptor becomes activated. [[Image:Domain_drawingnew.jpg|300px|left|thumb|Figure 2: The Two Domain Model]] | It is hypothesized that amylin binds to the receptor via a two-domain model (Figure 2). The model suggests a series of steps for how amylin binds. First, the c-terminus of amylin binds to the n terminus of the extracellular domain of the receptor. This binding factors the alignment of amylin's n-terminus to the primary GPCR binding site. Once both the c-terminus and n-terminus of amylin are bound, the receptor becomes activated. [[Image:Domain_drawingnew.jpg|300px|left|thumb|Figure 2: The Two Domain Model]] | ||
====RAMP-CTR Interface==== | ====RAMP-CTR Interface==== | ||
<scene name='10/1038828/Ramp_ctr_interface/ | <scene name='10/1038828/Ramp_ctr_interface/10'>RAMP CTR Interface </scene> is a key interaction that stabilizes the protein complex and positions the receptor to favorably bind to amylin. The RAMP-CTR interface extends into the plasma membrane, providing additional non-covalent bonding between the protein complex and the cell membrane. | ||
==== Extracellular Domain - RAMP interactions ==== | ==== Extracellular Domain - RAMP interactions ==== | ||
The extracellular domain of the CTR primarily contains polar residues in the extracellular space. In order to orient these residues to facilitate amylin binding, <scene name='10/1038828/Ctr_ramp_ecd_stablization/ | The extracellular domain of the CTR primarily contains polar residues in the extracellular space. In order to orient these residues to facilitate amylin binding, <scene name='10/1038828/Ctr_ramp_ecd_stablization/8'>RAMP makes hydrogen bonds with the CTR</scene> to increase the rigidity of the receptor binding site. | ||
== Clinical Significance == | == Clinical Significance == | ||
====Drug Development==== | ====Drug Development==== | ||
[[Image:align.png|300px|left|thumb|Figure 3:Amylin (green) aligned with Pramlintide (red)]] [https://en.wikipedia.org/wiki/Pramlintide Pramlintide] is a synthetic analog of amylin (Figure 3) that is commonly used in accordance with mealtime [https://en.wikipedia.org/wiki/Insulin insulin] to help treat type 1 and 2 diabetic patients <ref name="Hay"/>. This drug binds to AMYR competitively, increasing the AMYR GPCR signaling. Increased action of the AMYR receptor has been shown to modestly lower HbA1c levels, which is often accompanied by weight loss <ref name="Hoogwerf">PMID: 18561511</ref>. Pramlintide binds with more affinity than amylin due to mutations from hydrophobic residues A29, S28, and S29 to proline (Figure 4). The proline residues increase the rigidity of the ligand by creating unfavorable phi and psi angles, which improves the ability of the ligand to bind AMYR. Pramlintide treatment has also been shown to consistently reduce [https://en.wikipedia.org/wiki/Amyloid_plaques Amyloid β plaque] aggregation in rodent models with [https://en.wikipedia.org/wiki/Alzheimer%27s_disease Alzheimer’s disease] <ref name="Gingell">PMID:24169554</ref>. | [[Image:align.png|300px|left|thumb|Figure 3:Amylin (green) aligned with Pramlintide (red)]] [https://en.wikipedia.org/wiki/Pramlintide Pramlintide] is a synthetic analog of amylin (Figure 3) that is commonly used in accordance with mealtime [https://en.wikipedia.org/wiki/Insulin insulin] to help treat type 1 and 2 diabetic patients <ref name="Hay"/>. This drug binds to AMYR competitively, increasing the AMYR GPCR signaling. Increased action of the AMYR receptor has been shown to modestly lower HbA1c levels, which is often accompanied by weight loss <ref name="Hoogwerf">PMID: 18561511</ref>. Pramlintide binds with more affinity than amylin due to mutations from hydrophobic residues A29, S28, and S29 to proline (Figure 4). The proline residues increase the rigidity of the ligand by creating unfavorable phi and psi angles, which improves the ability of the ligand to bind AMYR. Pramlintide treatment has also been shown to consistently reduce [https://en.wikipedia.org/wiki/Amyloid_plaques Amyloid β plaque] aggregation in rodent models with [https://en.wikipedia.org/wiki/Alzheimer%27s_disease Alzheimer’s disease] <ref name="Gingell">PMID:24169554</ref>. | ||
[[Image:pram sequence align.png|300px|right|thumb|Figure 4:Pramlintide Sequence alignment with varying forms of amylin. Atoms C2—C7 and K1 of the N-terminal region are conserved. Y37 and T36 of the C-terminal region are also conserved.]] | [[Image:pram sequence align.png|300px|right|thumb|Figure 4: Pramlintide Sequence alignment with varying forms of amylin<ref name="Hay"/>. Atoms C2—C7 and K1 of the N-terminal region are conserved. Y37 and T36 of the C-terminal region are also conserved.]] | ||
It has been thought that [https://en.wikipedia.org/wiki/Missense_mutation missense mutations] in residues C2 and C7 of the amylin peptide could lead to an increased risk of Alzheimer's Disease <ref name="Grizzanti">PMID: 30282360</ref>. Because of the rigidity these cysteine resides provide, reductions of their disulfide interaction leads to an increased risk of amyloid plaques due to amylin misfolding and forming aggregates. During drug design, pharmaceutical companies have focused on maintaining amylin residues C2 and C7, as well as K1, which forms a hydrogen bond donor for the <scene name='10/1038828/N_term_disulfidenew/1'>E294 | It has been thought that [https://en.wikipedia.org/wiki/Missense_mutation missense mutations] in residues C2 and C7 of the amylin peptide could lead to an increased risk of Alzheimer's Disease <ref name="Grizzanti">PMID: 30282360</ref>. Because of the rigidity these cysteine resides provide, reductions of their disulfide interaction leads to an increased risk of amyloid plaques due to amylin misfolding and forming aggregates. During drug design, pharmaceutical companies have focused on maintaining amylin residues C2 and C7, as well as K1, which forms a hydrogen bond donor for the <scene name='10/1038828/N_term_disulfidenew/1'>E294 side chain</scene> and main chain carbonyl. Additionally, pharmaceutical companies have also opted to maintain residues <scene name='10/1038819/Amidated_c_term/9'>Y37 and T36</scene>, which are critical residues in stabilizing the C terminus of amylin to the receptor binding site. While there are hardly any differences in the helical portion of amylin and the synthetic analogue pramlintide, there is a difference in the extended random coil at the C terminus. | ||
In order to restore basal amylin levels in mice, researchers have performed PEGylation, the addition of polyethylene glycol, to amylin within residues 1-11 of the peptide, most likely at the two amine groups of K1<ref name="Guerreiro">PMID: 23818080</ref>. Administration of the modified amylin in mice showed evidence of reduced glycemia and prolonged action compared to endogenous amylin <ref name="Guerreiro"/>. | In order to restore basal amylin levels in mice, researchers have performed PEGylation, the addition of polyethylene glycol, to amylin within residues 1-11 of the peptide, most likely at the two amine groups of K1<ref name="Guerreiro">PMID: 23818080</ref>. Administration of the modified amylin in mice showed evidence of reduced glycemia and prolonged action compared to endogenous amylin <ref name="Guerreiro"/>. | ||