Heat shock Proteins (HSPs)

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Heat shock Proteins (HSPs)

• Heat shock proteins (HSP) are expressed in
  response to various biological stresses,
  including heat, high pressures, and toxic
  compounds. It is also one of the most abundant
  cellular proteins found under non-stress
  conditions
• Hsp90 is part of a family of proteins known as
  "chaperones," which are solely dedicated to
  helping other proteins fold and assume their
  proper functions.
• The chaperones Hsp70 and Hsp90 together with
  co-chaperones function to fold proteins in the
  cytoplasm. Sometimes Hsp70 and Hsp90 function
  sequentially to fold the same protein
• Cells are vigilant about getting these folds
  right because misfolded proteins can change the
  normal life of the cell. In some cases change is
  good, in others deadly.
• When HSP90 is compromised the number of
  morphological changes increases, which lead to
  formation of inactive or abnormally active
  polypeptides.

Domain structure of HSP90.
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Mad Cows, People yeast

• What do "mad cows," people with neurodegenerative
  diseases and an unusual type of yeast have in
  common? They are all experiencing the effects of
  misfolded proteins, according to HHMI
  investigator Susan Lindquist of the University of
  Chicago.
• Her research identified a role for HSP90 in the
  process of evolution.
• They have reported that fruit flies that make too
  little of the Hsp90 protein develop dramatic
  deformities, such as an extra antenna, additional
  bristles, notched wings or malformed eyes.
• The defects result from multiple hidden
  variations in the genome.
• When affected flies are interbred, these factors
  are enriched and subsequent generations have the
  same deformities, even though they have normal
  levels of Hsp90.

Susan LindquistUniversity of Chicago"My view
is that molecular chaperones are a way of
changing the traits of an organism that arose
very early in evolution. They might be as old as
RNA and DNA."  
Sourcehttp//www.hhmi.org/annual98/research/madco
w.html
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HSPs in protein folding

• The diagram shows the role of heat-shock proteins
  and a chaperonin in protein folding.  As the
  ribosome moves along the molecule of messenger
  RNA, a chain of amino acids is built up to form a
  new protein molecule.  The chain is protected
  against unwanted interactions with other
  cytoplasmic molecules by heat-shock proteins and
  a chaperonin molecule until it has successfully
  completed its folding.  

Source (http//www.cs.stedwards.edu/chem/Chemistr
y/CHEM43/CHEM43/HSP/FUNCTION.HTML)
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HSP90-alpha protein sequence

• HSP90alpha
  732 aa  HSP90beta
  724 aa  
• gtHSP90alphaMPEETQTQDQPMEEEEVETFAFQAEIAQLMSLIINTFY
  SNKEIFLRELISNSSDALDKIRYESLTDPSKLDSGKELHINLIPNKQDRT
  LTIVDTGIGMTKADLINNLGTIAKSGTKAFMEALQAGADISMIGQFGVGF
  YSAYLVAEKVTVITKHNDDEQYAWESSAGGSFTVRTDTGEPMGRGTKVIL
  HLKEDQTEYLEERRIKEIVKKHSQFIGYPITLFVEKERDKEVSDDEAEEK
  EDKEEEKEKEEKESEDKPEIEDVGSDEEEEKKDGDKKKKKKIKEKYIDQE
  ELNKTKPIWTRNPDDITNEEYGEFYKSLTNDWEDHLAVKHFSVEGQLEFR
  ALLFVPRRAPFDLFENRKKKNNIKLYVRRVFIMDNCEELIPEYLNFIRGV
  VDSEDLPLNISREMLQQSKILKVIRKNLVKKCLELFTELAEDKENYKKFY
  EQFSKNIKLGIHEDSQNRKKLSELLRYYTSASGDEMVSLKDYCTRMKENQ
  KHIYYITGETKDQVANSAFVERLRKHGLEVIYMIEPIDEYCVQQLKEFEG
  KTLVSVTKEGLELPEDEEEKKKQEEKKTKFENLCKIMKDILEKKVEKVVV
  SNRLVTSPCCIVTSTYGWTANMERIMKAQALRDNSTMGYMAAKKHLEINP
  DHSIIETLRQKAEADKNDKSVKDLVILLYETALLSSGFSLEDPQTHANRI
  YRMIKLGLGIDEDDPTADDTSAAVTEEMPPLEGDDDTSRMEEVD
  Green bases Geldanamycin-Binding Domain
• Brown bases Transmembrane segments as
  predicted by Tmap.
• Hydrophobic A,C, I,L, M, F, V,P, Y,W
  
• Hydrophilic R,N, H,D, E,Q, K, S,T Either G

GREASE output
TMAP output
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BLASTp

• With the HSP90 sequence in hand we used Blastp to
  find homologous sequences
• We were surprised to find a lot of homologous
  sequences across many species like Humans,
  Chicken,Pig, Mouse,Horse,Fish, Coral,fruit fly,
  mosquito, nematode, even crops like rice, maize
  tobacco.
• The first 100 matches had e-values ranging from 0
  to e-153, so they were very strong matches
  indicating a high degree of conservation of the
  protein through evolution.

ID Name Score Evalue 304882 heat shock 90kDa
protein 1, alpha Homo sapiens N...
1247 0.0 352285 heat shock protein 1, alpha Mus
musculus NP_0346... 825 0.0 761972 heat shock
protein 86 Rattus norvegicus NP_78693...
825 0.0 341493 heat shock protein 90A
Cricetulus griseus AAA369... 817 0.0 609431 hea
t shock protein 90 - chicken 816 0.0 609432 heat
shock protein 84 - mouse 745 0.0 449511 (Q9W6K6
) Heat shock protein hsp90 beta Salmo sala...
731 0.0 459017 heat shock protein hsp90
Oncorhynchus tshawytscha... 730 0.0 446434 heat
shock protein hsp90beta Danio rerio AAC2156...
729 0.0 361999 heat shock protein 90 Rattus
sp. AAB23369.1 S45... 724 0.0 460597 heat
shock protein 90 Pleurodeles waltl AAA92343...
719 0.0 738604 90-kDa heat shock protein Bombyx
mori BAB41209.1... 712 0.0 146263 Heat shock
protein 83 CG1242-PA Drosophila melano...
669 0.0 755572 heat shock protein 90
Dendronephthya klunzingeri...
662 0.0 226533 (P33126) Heat shock protein 82
Oryza sativa (Rice) 612 e-174 1888761 heat
shock protein 82 - common tobacco (fragment)
612 e-174 252633 heat shock protein Arabidopsis
thaliana CAA72513... 600 e-170 236351 (Q9XGF1)
HSP80-2 Triticum aestivum (Wheat)
598 e-169 283559 (Q08277) Heat shock protein 82
Zea mays (Maize) 593 e-168 152674 heat shock
protein 86 Plasmodium falciparum AAA6...
591 e-167 1899880 (Q8LLI6) Heat shock protein
Hsp90 Achlya ambisex... 579 e-164 245912 heat
shock protein 90 Lycopersicon esculentum AA...
544 e-153
 
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Multiple sequence alignment

• Multiple sequence alignments were done using
  ClustalW using different species the following
  unrooted phylogenetic tree was generated.

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• Rooted phylogenetic tree.

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Structure analysis

• We found open and closed conformations for
  the Geldanamycin-Binding Domain of the Human
  Hsp90 protein decided to study their
  differences. The parts in yellow are the
  selected residues which are also the
  transmembrane segments. The residues gtia in
  the sequence viewer show where the structures
  differ. The same region is depicted as the little
  grey segment at one end of the transmembrane
  segments

Comparison between Open Closed conformations of
human HSP90 alpha Transmembrane segments in yellow
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Comparison of HSP90 structures in Yeast Human

• We compared the HSP90 structures in yeast human
  found that the protein structures were very
  similar. The picture below shows the 2 structures
  superimposed with the highlighted portion showing
  an additional residue in the yeast sequence.
• Structures compared
•  1YES Human Hsp90 Geldanamycin-Binding Domain,
  "open" Conformation mmdbId7483
• 1AMW Atp Binding Site In The Hsp90 Molecular
  Chaperone,Saccharomyces cerevisiae (yeast)
  mmdbId7950

Red identical residues Blue similar
residues Yellow selected residues
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Microarray Data

• We found microarray data on the Cancer Genome
  Anatomy Project web site
• http//cgap.nci.nih.gov/Genes/GeneFinder

HSP90-alpha This screen shot shows the microarray
data for HSP90 alpha expression in various types
of cancers.
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HSP90-beta This screen shot shows the microarray
data for HSP90 beta expression in various types
of cancers.
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Current Studies on HSP90

• Changes in protein conformation are involved in
  some of the most devastating and intractable
  diseases.
• Studies in yeast may help us decipher the
  fundamental nature of these disorders, including
  Creutzfeldt-Jakob, Alzheimers, Huntingtons, and
  Parkinsons disease in humans and mad cow disease
  in cattle. Several of the protein culprits are
  being imported into yeast, which allows for the
  manipulation study of their folding transitions
  testing of therapeutic strategies.
• (http//www.wi.mit.edu/nap/pdfs/Directors_Report/d
  ir_lindquist02.pdf) 
• Conclusion
•  HSP90 is a powerful evolutionary mechanism that
  ensures apparent genetic stability at
  physiological conditions and at the same time
  allows the mutations that could rapidly become
  manifest under stress.     

• References
• http//www.stanford.edu/class/gene211/hsp90_search
  
• http//www.chemie.tu-muenchen.de/biotech/en/hsp90-
  e.html
• http//www.hhmi.org/annual98/research/madcow.html
• www.ashland.edu/kstine/Research/Stress20proteins
  .pdf