Insulin Signaling A Molecular View Shuchismita Dutta Ph

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Insulin Signaling: A Molecular View Shuchismita Dutta, Ph. D. Developed as part of the

Insulin Signaling: A Molecular View Shuchismita Dutta, Ph. D. Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Learning Objectives • Energy in Biology • Insulin Signaling • Insulin Resistance and Type

Learning Objectives • Energy in Biology • Insulin Signaling • Insulin Resistance and Type 2 Diabetes Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Learning Objectives • Energy in Biology • Insulin Signaling • Insulin Resistance and Type

Learning Objectives • Energy in Biology • Insulin Signaling • Insulin Resistance and Type 2 Diabetes Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Energy in Biology • ATP: Energy currency of cells • Main sources of ATP

Energy in Biology • ATP: Energy currency of cells • Main sources of ATP Glycogen Amino acids; Glycerol etc. ----Adenine-----Adenosine-------Adenosine monophosphate(AMP) ----Adenosine diphosphate(ADP)------Adenosine triphosphate(ATP)-------- Substrate Product Metabolic Process Glucose Pyruvate Glycolysis Pyruvate Acetyl Co. A Pyruvate Dehydrogenase Acetyl Co. A CO 2 Kreb’s Cycle Free fatty acids Acetyl Co. A Beta oxidation Fats Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Glucose Homeostasis • Pancreas produces – Insulin • Produced by b cells • Promotes

Glucose Homeostasis • Pancreas produces – Insulin • Produced by b cells • Promotes uptake of glucose from plasma – Glucagon • Produced by a cells • Promotes processes to release glucose into plasma Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Balancing Insulin and Glucagon Incretins GLP-1; GIP (Gut cells) Insulin production – High blood

Balancing Insulin and Glucagon Incretins GLP-1; GIP (Gut cells) Insulin production – High blood glucose – Glucagon – Incretin hormones • Glucose-dependent insulinotropic peptide (GIP) • Glucagon-like-peptide 1 (GLP-1) Glucagon production Insulin (b cells) Glucagon (a cells) – Low blood glucose – Insulin Developed as part of the RCSB Collaborative Curriculum Development Program 2016 Somatostatin (d cells)

Glucose Transporters 14 Types known; GLUT 1 -4 characterized • GLUT 1 • –

Glucose Transporters 14 Types known; GLUT 1 -4 characterized • GLUT 1 • – Main transporter in red blood cells and blood-tissue barriers – Ubiquitous; medium affinity – For basal-level glucose uptake • Glucose Co-transporters – Sodium Glucose co-transporters (SGLTs) GLUT 2 – Allows uptake and efflux of glucose in response to fed or fasted state – Mediates transport in liver, intestinal, kidney and pancreatic cells – Low affinity • GLUT 3 – Primary function in neurons, and circulating white blood cells – High affinity • GLUT 4 – Responsive to insulin – In adipocytes and muscle (skeletal and cardiac) cells http: //www. ncbi. nlm. nih. gov/pmc/articles/PMC 4104978/figure/F 6/ Developed as part of the RCSB Collaborative Curriculum Development Program 2016

http: //www. nature. com/nature/journal/v 526/n 7573/full/nature 14655. html Transporter GLUT 3 in Action Movie

http: //www. nature. com/nature/journal/v 526/n 7573/full/nature 14655. html Transporter GLUT 3 in Action Movie at http: //www. nature. com/nat ure/journal/v 526/n 7573/fig_ tab/nature 14655_SV 3. html Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Learning Objectives • Energy in Biology • Insulin Signaling • Insulin Resistance and Type

Learning Objectives • Energy in Biology • Insulin Signaling • Insulin Resistance and Type 2 Diabetes Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Insulin Signaling Insulin Receptor Insulin-Insulin Receptor Complex Extra-cellular Transmembrane Cytoplasmic http: //pdb 101. rcsb.

Insulin Signaling Insulin Receptor Insulin-Insulin Receptor Complex Extra-cellular Transmembrane Cytoplasmic http: //pdb 101. rcsb. org/motm/182 Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Insulin Receptor L 2 Fn. III-1 L 1 Fn. III-2 a. CT CR Extra-cellular

Insulin Receptor L 2 Fn. III-1 L 1 Fn. III-2 a. CT CR Extra-cellular Fn. III-3 Transmembrane Cytoplasmic http: //pdb 101. rcsb. org/motm/182 http: //www. ncbi. nlm. nih. gov/pmc/articl es/PMC 3793637/ Developed as part of the RCSB Collaborative Curriculum Development Program 2016 Ecto IR, PDB ID 3 loh

Insulin Binding to Receptor Insulin-Insulin Receptor Complex Ins: m. IR complex, PDB ID 3

Insulin Binding to Receptor Insulin-Insulin Receptor Complex Ins: m. IR complex, PDB ID 3 w 14 http: //pdb 101. rcsb. org/motm/182 a-CT of second IR protein interacts with insulin and first IR molecule Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Insulin Signal Transduction • IR Tyrosine Kinase domain: – Inactive state (PDB ID 1

Insulin Signal Transduction • IR Tyrosine Kinase domain: – Inactive state (PDB ID 1 irk) a mobile loop blocks entry of substrate – Active state (PDB ID 1 ir 3) several tyrosines on this loop are phosphorylated, loop swings out of active site, allowing ATP and other signaling proteins to bind. – Active site uses ATP to phosphorylate its targets. Tyrosine Kinase domain of IR showing inactive (PDB ID 1 IRK) and active (PDB IDs 1 IR 3) forms

Insulin Signaling 1. Insulin binds 6. Protein Synthesis 7. Cell Survival 8. Proliferation 2.

Insulin Signaling 1. Insulin binds 6. Protein Synthesis 7. Cell Survival 8. Proliferation 2. Tyrosine Kinase activation 3. Signaling molecules: IRS, PI 3 K, PDK 1, AKT, AS 160 etc. 4. Translocate GLUT 4 transporters 5. Glucose uptake Metabolism Glycolysis ATP + Pyruvate Lipogenesis Lipids Developed as part of the RCSB Collaborative Curriculum Development Program 2016 Storage Glycogenesis glycogen

Insulin Signal Transduction P PIP 2 IRS PI 3 K PIP 3 P PKC

Insulin Signal Transduction P PIP 2 IRS PI 3 K PIP 3 P PKC PDK 1 Glycogen Synthesis P GSK 3 Gluconeogenesis FOXO 1 Protein Synthesis m. TORC 1 AKT 2 Translocate to Glucose uptake P AS 160 P m. TORC 2 Developed as part of the RCSB Collaborative Curriculum Development Program 2016 GLUT 4 vesicle

Insulin Signal Transduction P PTP 1 B Glycogen Synthesis PIP 2 IRS PI 3

Insulin Signal Transduction P PTP 1 B Glycogen Synthesis PIP 2 IRS PI 3 K PIP 3 P PTEN PKC PDK 1 P GSK 3 Glyconeogenesis FOXO 1 Protein Synthesis m. TORC 1 AKT 2 Translocate to Glucose uptake P AS 160 P m. TORC 2 Developed as part of the RCSB Collaborative Curriculum Development Program 2016 GLUT 4 vesicle

Understanding AKT P AKT 2 P • N-terminal PH domain localizes AKT to plasma

Understanding AKT P AKT 2 P • N-terminal PH domain localizes AKT to plasma membrane ( levels of PI(3, 4, 5)P 3 or PI(3, 4)P 2) • Catalytic domain activated by phosphorylation of Thr – T 309 by PDK and S 474 by m. TORC 1 AKT Kinase domain showing inactive (PDB ID 1 MRV) and active (PDB IDs 1 O 6 L) forms

Learning Objectives • Energy in Biology • Insulin Signaling • Insulin Resistance and Type

Learning Objectives • Energy in Biology • Insulin Signaling • Insulin Resistance and Type 2 Diabetes Developed as part of the RCSB Collaborative Curriculum Development Program 2016

Insulin Resistance • Insulin is present but signaling is disrupted at the level of

Insulin Resistance • Insulin is present but signaling is disrupted at the level of IRS, AKT, AS 160, etc. • Body tries to compensate by making more insulin P PIP 2 IRS PI 3 K PIP 3 P PKC Glucose uptake PDK 1 Glycogen Synthesis P GSK 3 Glyconeogenesis FOXO 1 Protein Synthesis m. TOR AKT 2 P AS 160 P m. TOR/ Developed as part of the RCSB Collaborative Curriculum Development Program 2016 GLUT 4 vesicle

A Case of Insulin Resistance • A family with severe insulin resistance and diabetes

A Case of Insulin Resistance • A family with severe insulin resistance and diabetes due to a mutation in AKT 2 Science. 2004 May 28; 304: 1325 -8. • Molecular Exploration … (see hands-on exercise attached)

What May Happen in R 274 H Active Site Residues R 274 Substrate peptide

What May Happen in R 274 H Active Site Residues R 274 Substrate peptide PDB ID 1 o 6 l, Yang et al. , 2002 TPO 309 Mutation of Arg 274 to His would disrupt the H-bonds holding the activation loop in the active state

Summary • Energy in Biology – ATP is energy currency • Insulin Signaling –

Summary • Energy in Biology – ATP is energy currency • Insulin Signaling – Reception, Signal Transduction and Response • Insulin Resistance and Type 2 Diabetes – Mutations may disrupt proteins in Insulin signaling Developed as part of the RCSB Collaborative Curriculum Development Program 2016