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CA García Sepúlveda MD PhD

Chaperones. CA García Sepúlveda MD PhD. Laboratorio de Genómica Viral y Humana Facultad de Medicina, Universidad Autónoma de San Luis Potosí. Chaperones.

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CA García Sepúlveda MD PhD

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  1. Chaperones. CA García Sepúlveda MD PhD Laboratorio de Genómica Viral y HumanaFacultad de Medicina, Universidad Autónoma de San Luis Potosí

  2. Chaperones The reactivity of protein surfacesmeans that incorrect interactionsmay occur unless the process of folding a protein after translationis controlled. Some proteins are able to acquire their mature conformation spontaneously =Self assembly. A test for this ability = denature the protein and determine whether it can then renature into the active form → implies that the internal interactions are intrinsically directed toward the right conformation. https://www.youtube.com/watch?v=mretwWl5BYI

  3. Chaperones • When this does not happen, and alternative sets of interactions can occur, a protein may become trapped in a stable conformation THAT IS NOT THE INTENDED FINAL FORM !

  4. Chaperones • Proteins in this category cannot self-assemble. Their acquisition of proper structure requires the assistance of achaperone. • Chaperones are proteins that mediate correct assembly by causing a target protein to acquire one possible conformation instead of others. • Accomplished by binding to reactive surfaces in the target protein that are exposed during the assembly process and preventing them from interacting. • Chaperones function by preventing formation of incorrect structures rather than by promoting formation of correct structures.

  5. Chaperones 1.- Initial acquisition of the correct conformation. • The ability of chaperones to recognize incorrect protein conformations allows them to play two related roles concerned with protein structure: • As a protein is synthesized (as it exits the ribosome) it is in an unfolded form. • Spontaneous folding then occurs as the emerging sequence interacts with regions of the protein that were synthesized previously. • Chaperones influence the folding process by controlling the accessibility of the reactive surfaces.

  6. Chaperones 2.- Asisting renaturation or terminal degradation. • When a protein is denatured, new regions are exposed and become able to interact. • These interactions are similar to those that occur when a protein (transiently) misfolds as it is initially synthesized. • They are recognized by chaperones as comprising incorrect folds. • Chaperones assist renaturation or degradation.

  7. Chaperones 1.- Initial acquisition of the correct conformation. • The ability of chaperones to recognize incorrect protein conformations allows them to play two related roles concerned with protein structure: https://www.youtube.com/watch?v=d1QIEQEyYRo

  8. Chaperones • Chaperones also required to assist the formation of oligomeric structures and for the transport of proteins through membranes. • Protein folding is an important feature of membrane passage. • It may be necessary to maintain a protein in an unfolded state because of the geometry of passage. • Chaperones may prevent a protein from acquiring a conformation that would prevent passage through the membrane. • Once the protein has passed through the membrane, it may require another chaperone to assist with folding to its mature conformation.

  9. Chaperones • Two major groups of chaperones have been well characterized: • The Hsp70 system (Hsp70, Hsp40 & GrpE). • The chaperonin system (Hsp10, Hsp60) Prokaryotes Eukaryotes

  10. Chaperones Eukaryotes • Two major groups of chaperones have been well characterized: • The Hsp70 system (Hsp70, Hsp40 & GrpE). Prokaryotes • Functions on: • newly synthesized protein • proteins being transported through membranes • proteins denatured by stress. The name reflects their induction by heat shock.

  11. Chaperones • Two major groups of chaperones have been well characterized: • The chaperonin system (Hsp10, Hsp60) Consists of a large oligomeric assembly. Also minimize the damage caused to proteins by heat denaturation.

  12. The Hsp70 System • The Hsp70 system (Hsp70, Hsp40 & GrpE). • The Hsp70 family is found in bacteria, eukaryotic cytosol, in the ER & in chloroplasts/mitochondria. • Members help protein folding by multiple cycles of association and dissociation to nascent peptide. • Hsp70 functions in conjunction with two further components,best characterized in bacteria as DnaJ and GrpE. Prokaryotes Eukaryotes

  13. The Hsp70 System • Different members of the Hsp70 class function on different types of targetproteins. • Cytosolic Hsp70 & Hsc70 act on nascent proteins on free ribosomes.

  14. The Hsp70 System • Different members of the Hsp70 class function on different types of targetproteins. • ER, mitochondrial/choloroplast variants BiP,Grp78 & Kar2 act on nascent peptides as they emerge into the interior of the organelle on passing through the membrane.

  15. The Hsp70 System • Hsp40 (DnaJ) binds to a nascent protein as it emerges from the ribosome. • Hsp40 contains a region called the J domain which interacts with Hsp70. Hsp40 (DnaJ)

  16. The Hsp70 System • Hsp70 (DnaK) binds to both Hsp40 and to the unfolded protein. Hsp70 (DnaK)

  17. The Hsp70 System • The interaction of Hsp70 with Hsp40 stimulates Hsp70 ATPase activity. ATPase

  18. The Hsp70 System • The ADP-bound form of the complex remains associated with the protein substrate until GrpE displaces the ADP. GrpE displaces ADP

  19. The Hsp70 System • This causes dissociation of the complex: • Hsp40 (DnaJ) dissociates • ATP binds to Hsp70 (DnaK) • Hsp70 dissociates. Complex dissociation

  20. The Hsp70 System • As the protein chain lengthens, Hsp70 (DnaK) may dissociate from one binding site and reassociate with another. • Thus releasing parts of the substrate protein to fold in an ordered manner. • Finally, the intact protein is released from the ribosome, folded into its mature conformation Protein release

  21. The Chaperonin System • Large (oligomeric) structure with a hollow cavity used for folding or degradation of proteins. • The typical structure is a ring of many subunits, forming a cylinder. Target protein is placed in a controlledenvironment (cavity). • In folding, the closed environment • prevents the target protein from wrong • interactions with other proteins. • In the case of degradation, isolation • makes for a more controlled process • than would be possible in open cytosol. • The energy for these processes is provided by hydrolysis of ATP.

  22. The Chaperonin System • GroEL (prokaryote) forms an oligomer of two rings, each comprising a hollow cylinder made of 7 subunits. • The central hole (4.5 nm) is blocked at the equator of each ring by the COOH ends of the subunits, which protrude into the interior. • This structure associateswith a heptamer formed of subunits of Hsp10 (GroES in E. coli). • The dome caps one opening of the cylinder. NOTES: 1.- Entire structure has a mass ~10x106 daltons. 2.- Comparable to a small ribosomal subunit. 3.- GroEL sometimes called a chaperonin & GroES a co-chaperonin.

  23. The Overview (iBiology with Arthur Horwich) https://www.youtube.com/watch?v=jOhNyVjkChM

  24. die Proteinfaltung auf Max Planck Gesselschaft https://www.youtube.com/watch?v=nEHe3Aie9Ek

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