1. Small Heat Shock Proteins
Meagen Bailey

2. HSP- Overview/Structure
Classified into 6 structural classes
- HSP100, HSP90, HSP70, HSP60, sHSP and ubiquitin
Involved in folding of denatured proteins
Respond/elevated in temp and stress changes
Heat shock factors (HSF) and heat-shock elements (HSEs)
Associated with nuclei, cytoskeleton and membranes
HSP ->are classified into 6 structural conserved classes according to their molecular weight
Synthesized in dramatically increased amounts after a brief exposure of cells to an elevated temperature (Ex. 42°C for cell that normally live at 37°C)
High temperatures and other stresses, such as altered pH and oxygen deprivation make it more difficult for proteins to form their proper structures and cause some already structured proteins to unfold. They bind partially to denatured proteins, preventing irreversible protein aggregation during stress.
Transcriptional regulation of sHSPs depends on particular activation of heat shock factors (HSF) which recognize the highly conserved heat-shock elements (HSEs). After the heat stress has been released, the sHSPs are quite stable, suggesting that sHSPs may be important for recovery as well.

3. Overview- General Structure
α-crystallin
kDa
-αA-crystallin & αB-crystallin
amino acid sequences=> α-crystallin core domain
Forms distinct structural and functional unit
N terminus => variable in sequence and length
C terminus=> conveys stability and solubility
α-crystallin, ranges in mass from 12 to 43 kDa, a major lens protein that is composed of two similar subunits-> α-crystallin and αB-crystallin, that form large complexes and functions as molecular chaperones
sHSPs are characterized by an amino acid sequence known as the α-crystallin core, that is located in the C-terminal domain
Info Summary:
All members of the family are characterized by the presence of a homologous sequence of about 80 residues, which has been dubbed the “α-crystallin domain” and probably forms a distinct structural and functional unit. This domain is preceded by an N-terminal region of variable length, which shows little of no similarity between the various branches of the family. A short and variable sequence, but containing a conserved motif, extends C-terminally from the “α-crystallin domain”.

5. Primary Structure
Picture:
Schematic and linear representation of the domain structure and co-chaperone-binding sites of Hsp70 and Hsp90. The amino-acid residue numbers, domains and co-chaperone-binding sites are indicated. (A) Domain structure of human Hsp70. The C-terminal EEVD motif is characteristic for cytosolic Hsp70s and is involved in binding of TPR-domain-containing proteins. (B) Domain structure of human Hsp90α. The C-terminal MEEVD motif is involved in binding of TPR-domain-containing proteins. Binding sites of co-chaperones on Hsp90 are a composite of sites of interaction for mammalian and yeast Hsp90 [adapted from previous reports

6. Secondary Structure
2 hydrophobic β-sheet- rich motifs, connected by a hydrophobic α- helical region
α-helix
β-sheets
Info:
Picture:
Schematic of the SBD of DnaK. This secondary structure representation of the SBD of DnaK in a partial space-filling and ribbon representation is derived from the crystal structure of the C-terminal domain bound to the peptide NRLLLTG (shown in pink; Ref. 13 ). The SBD is composed of -pleated sheets comprising the peptide binding cleft and an α-helical lid region. An arch formed by residues M404 and A429 (gray) encloses the peptide backbone and a deep pocket formed by residue V436 (green) accommodates hydrophobic side chains.

7.
Cell Book (pg 390)
(A) Showing the catalysis of protein refolding. A misfolded protein is initially captured by hydrophobic interactions along one rim of the barrel. The subsequent binding of ATP plus a protein cap increases the diameter of the barrel rim, which may transiently stretch (partly unfold) the client protein. This also confines the protein in an enclosed space, where it has a new opportunity to fold. After about 15 seconds, ATP hydrolysis occurs, weakening the complex. Subsequent binding of another ATP molecule eject the protein, whether folded or not, and the cycle repeats.
(B) The structure of GroEL bound to its GroES cap, as determined by X-ray crystallography. On the left is shown the outside of the barrel-like structure and on the right a cross section through its center.

8. Tertiary Structure αA (67-101) αB (70-105)
Shares ~60% sequence homology
Aa sequence w/in this segment contain charged residues
Charged residues and ANS binding=> role in binding
Hydrophobic interaction
Info:
Bind to hydrophobic domains of unfolded proteins
Bind to nonpolar segments
Target unstructured stretch of hydophobic amino acids flanked by basic residues and lacking acid residues
Picture:

9. Quaternary Structure Depend on N- terminal (18-19 aa)
Mulitmeric complex of αA- and αB- crystallines
35-40 subunits
~800kDa molecules
Information Info:
Polydisperse quaternary structure appear, in part, depend upon the N-terminal 18 and 19 amino acids that are essential for subunit interaction in polydisperse sHsps.
Multimeric complexes…..
Picture Info:
Figure 2 Features of sHsp structures. (a) Quaternary structures of sHsps, determined either by cryo-EM or X-ray crystallography. Hsp26 from S. cerevisiae (24 subunits, cryo-EM; H. Saibil and H. White (Birkbeck College, London), personal communication), αB-crystallin from Homo sapiens (16 subunits, cryo-EM)17, Hsp16.5 from M. jannaschii (24 subunits, crystal)13, Hsp16.3/Acr1 from M. tuberculosis (12 subunits, cryo-EM)14 and Hsp16.9 from T. aestivum (12 subunits, crystal)16.(b,c) M. jannaschii Hsp16.5 (b), whose quaternary structure consists of 24
identical subunits arranged into a tetrahedral hollow sphere, andT. aestivum Hsp16.9 (c), a barrel-like assembly of 12 subunits; crosssections show inner surfaces of both proteins; colors distinguish neighboring dimers. (d,e) Crystal structures of two sHsp dimers, from M. jannaschii Hsp16.5 (d; PDB code 1SHS) and T. aestivum Hsp16.9 (e; PDB code1GME), highlighting three functionally important regions. Green, N-terminal region; blue, α-crystallin domain; magenta, C-terminal extension; gray, the other sHsp in the dimer; dotted line, N-terminal residues not resolved in the structures. (f) Domain organization of sHsps.

10. Small Heat Shock Proteins (sHSP) are chaperone-like proteins involved in multiple cellular processes such as denatured protein aggregate prevention, apoptose inhibition, or cytosqueleton protection. These proteins are large assemblies generally composed of several copies of the same monomer. We aim to understand the building process of these complexes, starting from the dynamical properties of the monomer (figure a), to the association into dimers (figure b) to finally reach the functional assembly (figure c). The study of these objects requires many bioinformatics tools such as multiple sequence alignments, homology modelling, molecular dynamics and docking simulation with a modelisation scale ranging from all-atom to coarse grain. Based on sequence informations, we are currently working on the structural properties of the different regions of the sHSP monomers, namely the Nter, Alpha Crystallin Domain (ACD) and Cter regions.

11. Small Heat Shock Proteins (sHSPs)
Marshall Aldrich

12. Overview Activation Functions Chaperone Activity Cell Development
Cell Protection

13. Activation

14. Chaperoning

15. Chaperoning
GroEL/GroES Cycle

16. Cell Development

17. Cell Protection
Protecting neurons from acute and chronic effects of alcohol

18. Cell Protection
αB-Crystallin prevents neuropathy and cataracts

19. Cell Protection Cont.

20. Works Cited
Cates, Jordan, Garrett C. Graham, Natalie Omattage, Elizabeth Pavesich, Ian Setliff, Jack Shaw, Caitlin Lee Smith, and Ovidiu Lipan. “Sensing the Heat Stress by Mammalian Cells." BMC Biophysics 4.1 (2011): 16. Web.
Clark, A.r., C.e. Naylor, C. Bagnéris, N.h. Keep, and C. Slingsby. "Crystal Structure of R120G Disease Mutant of Human αB-Crystallin Domain Dimer Shows Closure of a Groove." Journal of Molecular Biology (2011). Web.
Crippa, V., D. Sau, P. Rusmini, A. Boncoraglio, E. Onesto, E. Bolzoni, M. Galbiati, E. Fontana, M. Marino, S. Carra, C. Bendotti, S. De Biasi, and A. Poletti. "The Small Heat Shock Protein B8 (HspB8) Promotes Autophagic Removal of Misfolded Proteins Involved in Amyotrophic Lateral Sclerosis (ALS)." Human Molecular Genetics (2010): Web.
Houck, Scott A., and John I. Clark. "Dynamic Subunit Exchange and the Regulation of Microtubule Assembly by the Stress Response Protein Human αB Crystallin." Ed. Stefan Wölfl. PLoS ONE 5.7 (2010): E Web.
Kregel, Kevin C. "Invited Review: Heat Shock Proteins: Modifying Factors in Physiological Stress Responses and Acquired Thermotolerance." J Appl Physiol 92 (2002): Web.

21. Works Cited Cont.
Narberhaus, F. " -Crystallin-Type Heat Shock Proteins: Socializing Minichaperones in the Context of a Multichaperone Network." Microbiology and Molecular Biology Reviews 66.1 (2002): Web. Stengel, F., A. J. Baldwin, A. J. Painter, N. Jaya, E. Basha, L. E. Kay, E. Vierling, C. V. Robinson, and J. L. P. Benesch. "Quaternary Dynamics and Plasticity Underlie Small Heat Shock Protein Chaperone Function." Proceedings of the National Academy of Sciences (2010): Web. Toth, Melinda E., Szilvia Gonda, Laszlo Vigh, and Miklos Santha. "Neuroprotective Effect of Small Heat Shock Protein, Hsp27, after Acute and Chronic Alcohol Administration." Cell Stress Chaperones 15.6 (2010): Web. Voet, Donald, and Judith G. Voet. Biochemistry. New York: J. Wiley & Sons, Web. Yenari, Midori A. "Heat Shock Proteins and Neuroprotection." Madame Curie Bioscience Database (2000). Web. < NBK6614/>.