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F44E5.4 (Heat Shock Protein 70)

F44E5.4 (Heat Shock Protein 70). IMBBR 315. Christopher Ricupero. Clones. 58.3% of colonies picked vs. successful clones sequenced. Clones. 58.3% of colonies picked vs. successful clones sequenced. 14D6-57 Sanger BLAST N Results. Insert size – small ~ 418 bp Sequenced 1 direction

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F44E5.4 (Heat Shock Protein 70)

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  1. F44E5.4 (Heat Shock Protein 70) IMBBR 315 Christopher Ricupero

  2. Clones 58.3% of colonies picked vs. successful clones sequenced

  3. Clones 58.3% of colonies picked vs. successful clones sequenced

  4. 14D6-57SangerBLAST N Results • Insert size – small ~ 418 bp • Sequenced 1 direction • Spanned entire insert Exon Intron Exon

  5. 14D6-57SangerBLAST N Results • Insert size – small ~ 418 bp • Sequenced 1 direction • Spanned entire insert • Interesting Observation • Sequence matching two unique genomic regions, in two directions What is going on?

  6. Sequence Summary for: F44E5 • P(N) 8.6 e-65 -Sequence matches hsp70 (Heat Shock Protein) • Gene duplication • Protein - 645 AAs • RNAi - Wild type morphology What is this protein?

  7. Structure • Size – 70kD • Length – 645 AAs • ATP dependent • Single Monomer • 3 Domains • N –Terminal - ATP Binding and hydrolysis (44 kDa) Homologous to Actin and Hexokinase • Substrate Binding – Hydrophobic regions (15-20 kDa) • C-terminal –Lid for substrate binding (15 -20 kDa) • Allosteric properties – conformational changes What does it do?

  8. Hsp70 (Heat Shock Protein) Essential in Protein Folding Cellular stress protection • Key Points: • A Molecular chaperone • Inducible during “stressful conditions” (heat, toxins, etc..) • Ubiquitous expression - In all cells at all biological levels • Highly conserved family (Hsp 40, 60, 90 etc..) • Consists of stress-inducible and constitutive family members (Hsc 70, Hsp72)

  9. Hsp70 - Molecular chaperone • Recognizes & binds regions rich in hydrophobic residues • Stabilizes & prevents misfolding by open conformation • Prevents aggregation and degradation during synthesis • Multiple Hsp70 complexes per polypeptide chain • Releases or transfers to other chaperones/chaperonins (Hsp60, Hsp90)

  10. Hsp70 – Cellular Protection • Stress Induced –Prevents protein denaturation and incorrect polypeptide aggregation during exposure to physiochemical insults • Hsp70 expression linked in several different models of neurodegenerative diseases

  11. Hsp70 – Neurodegenerative Diseases Hsp 70 Fonte, Virginia et al. (2002) Proc. Natl. Acad. Sci. USA 99, 9439-9444 • C. Elegans model of Alzheimer’s disease • Hsp70 and Hsp16 expression patterns correlated with phenotype

  12. Hsp70 -Additional Functions Involved in various cellular functions and diseases: Apoptosis Clathrin cage disassembly Parkinson's Disease Dauer Stage Translocation Cancer Prions

  13. Acknowledgements Thank you! Dr. Marty Nemeroff Dr. Monica Driscoll Indrani Chatterjee Brian Gelfand The rest of 315 staff Questions?

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