Lecture 3 "Molecular Chaperones & Protein quality control"

they assist folding and help mantain protein homeostasis

• fomration of primary aa sequence, flexibility of peptide bonds and interactions of side chains

• formation of secondary, tertiary and quarternary structures

• the native form of a protein has thermodynamically most stable form

• protein can take native form in absence of other proteins/assistance

• the primary structure

• small proteins can fold to the native form by themself

• highly conserved throughout the whole kingdom

• most of them encoded by multigene families which named by their molecular size

• numerous members expressed under stressful conditions, most prominent after heat stress

  > Heat shock proteins (HSP) vs. heat shock cognate (HSC)

• members of each family mostly in cellular compartments

• recognize and binidng of unfolded/denatured proteins

• asissting, but not catalyzing protein folding

  > provide the right environment

• protein quality control

• mantainance of cellular protein homeostasis

  > check for correct transfer, native state tec.

• add radioactive methionine, it binds during protein synthesis and is easy to detect

• The cell uses a team of chaperones to help proteins fold correctly.

• Main helpers are Hsp70/DnaK, Hsp40/DnaJ, trigger factor, and GroEL/ES.

• Their jobs are to help folding, stop aggregation, and rescue stressed proteins.

• They do not build the final structure themselves; they just help the protein reach it.

• Main idea: many chaperones work together to keep proteins functional

• HSP90

• HSP70

• HSP60

• HSP20

• Hsp70 starts in an ATP-bound open state, where it can bind a misfolded or unfolded protein.

• DnaJ/Hsp40 brings the substrate to Hsp70 and stimulates ATP hydrolysis.

• After ATP is hydrolyzed to ADP, Hsp70 closes tightly around the protein.

• This helps hold the protein, prevent aggregation, and give it time to fold correctly.

• Nucleotide exchange factors then replace ADP with ATP, reopening Hsp70 and releasing the substrate so another cycle can begin

• stimulation of the ATPase activity of HSP70s by the J-domain

• J-domain recruits Hsp70 to substrates

• few j-proteins bind only one substrate

• some j-proteins have borad substrate specificity, with a breath similar to HSP70 intercations

• prevention of protein aggregation

• protein folding/refolding

• facilitating protein degradation and sequestration

• protein translocation across membranes

• remodeling of protein complexes

• Housekeeping activities

  • Cooperation with other protein folding and quality control machineries

  • De novo protein folding

  • Protein translocation across membranes

  • Assembly/dissasembly of protein complexes

  • Regulation of protein activity

  • Protection from proteolysis

• Stress related activities

  • Protein aggregation prevention

  • Protein disaggregation

  • Protein refolding

  • Protein degradation

• Two groups:

  1. bacterial cytosol (GroEL), mitochondria (HSP 60) and chloroplasts (Cpn 60)

  2. Group 2: In archea (thermosome) and eukaryotic cytosol

• chaperonin (HSP 60)

• Hop and Cdc delay ATP hydrolysis

• Aha1 stabilized NTD dimerizatioh > accelerating ATP hydrolysis

• ß23 stabilizes closed dimer > slower release of substrate protein

• contains C-terminal domain, which is important for the interaction with co-cahperones that contain TPR domains

• cell signaling molecules: kinased (e.g. insulin receptor)

• receptors: steroid, glucocorticoids

• TF: HS, Hypoxia induced factor

• cytoskeleton: actin, tubulin (protection from heat stress)

• Stress:

  • heat

  • oxidative

  • heavy metal

  • salt

• it can hide he effects of mutations by helping unstable proteins still function

• Holdase: sHsps bind unfolding proteins and keep them from aggregating.

• Sequestration: they trap damaged proteins in a safe complex.

• Protection / thermal buffering: they stabilize proteins during heat or stress.

• Handoff to refolding chaperones: later, the substrate can be passed to Hsp70/Hsp100 for recovery