Skip to main content
SpringerLink
Log in

Heterooligomeric complexes of human small heat shock proteins

  • Original Paper
  • Published:
Cell Stress and Chaperones Aims and scope

Abstract

Oligomeric association of human small heat shock proteins HspB1, HspB5, HspB6 and HspB8 was analyzed by means of size-exclusion chromatography, analytical ultracentrifugation and chemical cross-linking. Wild-type HspB1 and Cys mutants of HspB5, HspB6 and HspB8 containing a single Cys residue in position homologous to that of Cys137 of human HspB1 were able to generate heterodimers cross-linked by disulfide bond. Cross-linked heterodimers between HspB1/HspB5, HspB1/HspB6 and HspB5/HspB6 were easily produced upon mixing, whereas formation of any heterodimers with participation of HspB8 was significantly less efficient. The size of heterooligomers formed by HspB1/HspB6 and HspB5/HspB6 was different from the size of the corresponding homooligomers. Disulfide cross-linked homodimers of small heat shock proteins were unable to participate in heterooligomer formation. Thus, monomers can be involved in subunit exchange leading to heterooligomer formation and restriction of flexibility induced by disulfide cross-linking prevents subunit exchange.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10

Similar content being viewed by others

Abbreviations

DTT:

Dithiothreitol

ME:

Mercaptoethanol

SEC:

Size-exclusion chromatography

sHsp:

Small heat shock proteins

References

  • Aquilina JA, Benesch JL, Ding LL, Yaron O, Horwitz J, Robinson CV (2004) Phosphorylation of alphaB-crystallin alters chaperone function through loss of dimeric substructure. J Biol Chem 279(27):28675–28680

    Article  PubMed  CAS  Google Scholar 

  • Bagneris C, Bateman OA, Naylor CE, Cronin N, Boelens WC, Keep NH, Slingsby C (2009) Crystal structures of alpha-crystallin domain dimers of alphaB-crystallin and Hsp20. J Mol Biol 392(5):1242–1252

    Article  PubMed  CAS  Google Scholar 

  • Baldwin AJ, Hilton GR, Lioe H, Bagneris C, Benesch JL, Kay LE (2011a) Quaternary Dynamics of alphaB-Crystallin as a Direct Consequence of Localised Tertiary Fluctuations in the C-Terminus. J Mol Biol. doi:10.1016/j.jmb.2011.07.017

  • Baldwin AJ, Lioe H, Robinson CV, Kay LE, Benesch JL (2011b) alphaB-Crystallin Polydispersity Is a Consequence of Unbiased Quaternary Dynamics. J Mol Biol. doi:10.1016/j.jmb.2011.07.016

  • Baranova EV, Weeks SD, Beelen S, Bukach OV, Gusev NB, Strelkov SV (2011) Three-dimensional structure of alpha-crystallin domain dimers of human small heat shock proteins HSPB1 and HSPB6. J Mol Biol 411(1):110–122

    Article  PubMed  CAS  Google Scholar 

  • Benesch JL, Ayoub M, Robinson CV, Aquilina JA (2008) Small heat shock protein activity is regulated by variable oligomeric substructure. J Biol Chem 283(42):28513–28517

    Article  PubMed  CAS  Google Scholar 

  • Berengian AR, Parfenova M, McHaourab HS (1999) Site-directed spin labeling study of subunit interactions in the alpha-crystallin domain of small heat-shock proteins. Comparison of the oligomer symmetry in alphaA-crystallin, HSP 27, and HSP 16.3. J Biol Chem 274(10):6305–6314

    Article  PubMed  CAS  Google Scholar 

  • Bova MP, Ding LL, Horwitz J, Fung BK (1997) Subunit exchange of alphaA-crystallin. J Biol Chem 272(47):29511–29517

    Article  PubMed  CAS  Google Scholar 

  • Bova MP, McHaourab HS, Han Y, Fung BK (2000) Subunit exchange of small heat shock proteins. Analysis of oligomer formation of alphaA-crystallin and Hsp27 by fluorescence resonance energy transfer and site-directed truncations. J Biol Chem 275(2):1035–1042

    Article  PubMed  CAS  Google Scholar 

  • Bukach OV, Seit-Nebi AS, Marston SB, Gusev NB (2004) Some properties of human small heat shock protein Hsp20 (HspB6). Eur J Biochem 271(2):291–302

    Article  PubMed  CAS  Google Scholar 

  • Bukach OV, Glukhova AE, Seit-Nebi AS, Gusev NB (2009) Heterooligomeric complexes formed by human small heat shock proteins HspB1 (Hsp27) and HspB6 (Hsp20). Biochim Biophys Acta 1794(3):486–495

    PubMed  CAS  Google Scholar 

  • Carra S, Seguin SJ, Landry J (2008) HspB8 and Bag3: a new chaperone complex targeting misfolded proteins to macroautophagy. Autophagy 4(2):237–239

    PubMed  CAS  Google Scholar 

  • Datta SA, Rao CM (2000) Packing-induced conformational and functional changes in the subunits of alpha-crystallin. J Biol Chem 275(52):41004–41010

    Article  PubMed  CAS  Google Scholar 

  • den Engelsman J, Boros S, Dankers PY, Kamps B, Vree Egberts WT, Bode CS, Lane LA, Aquilina JA, Benesch JL, Robinson CV, de Jong WW, Boelens WC (2009) The small heat-shock proteins HSPB2 and HSPB3 form well-defined heterooligomers in a unique 3 to 1 subunit ratio. J Mol Biol 393(5):1022–1032

    Article  Google Scholar 

  • Fontaine JM, Rest JS, Welsh MJ, Benndorf R (2003) The sperm outer dense fiber protein is the 10th member of the superfamily of mammalian small stress proteins. Cell Stress Chaperones 8(1):62–69

    Article  PubMed  CAS  Google Scholar 

  • Fontaine JM, Sun X, Benndorf R, Welsh MJ (2005) Interactions of HSP22 (HSPB8) with HSP20, alphaB-crystallin, and HSPB3. Biochem Biophys Res Commun 337(3):1006–1011

    Article  PubMed  CAS  Google Scholar 

  • Fontaine JM, Sun X, Hoppe AD, Simon S, Vicart P, Welsh MJ, Benndorf R (2006) Abnormal small heat shock protein interactions involving neuropathy-associated HSP22 (HSPB8) mutants. FASEB J 20(12):2168–2170

    Article  PubMed  CAS  Google Scholar 

  • Fuchs M, Poirier DJ, Seguin SJ, Lambert H, Carra S, Charette SJ, Landry J (2010) Identification of the key structural motifs involved in HspB8/HspB6-Bag3 interaction. Biochem J 425(1):245–255

    Article  CAS  Google Scholar 

  • Haley DA, Bova MP, Huang QL, McHaourab HS, Stewart PL (2000) Small heat-shock protein structures reveal a continuum from symmetric to variable assemblies. J Mol Biol 298(2):261–272

    Article  PubMed  CAS  Google Scholar 

  • Haslbeck M, Franzmann T, Weinfurtner D, Buchner J (2005) Some like it hot: the structure and function of small heat-shock proteins. Nature Struct Mol Biol 12(10):842–846

    Article  CAS  Google Scholar 

  • Hayes D, Napoli V, Mazurkie A, Stafford WF, Graceffa P (2009) Phosphorylation dependence of hsp27 multimeric size and molecular chaperone function. J Biol Chem 284(28):18801–18807

    Article  PubMed  CAS  Google Scholar 

  • Horwitz J (2003) Alpha-crystallin. Exp Eye Res 76(2):145–153

    Article  PubMed  CAS  Google Scholar 

  • Huey KA, Thresher JS, Brophy CM, Roy RR (2004) Inactivity-induced modulation of Hsp20 and Hsp25 content in rat hindlimb muscles. Muscle Nerve 30(1):95–101

    Article  PubMed  CAS  Google Scholar 

  • Jehle S, Vollmar BS, Bardiaux B, Dove KK, Rajagopal P, Gonen T, Oschkinat H, Klevit RE (2011) N-terminal domain of {alpha}B-crystallin provides a conformational switch for multimerization and structural heterogeneity. Proc Natl Acad Sci U S A 108(16):6409–6414

    Article  PubMed  CAS  Google Scholar 

  • Kappe G, Franck E, Verschuure P, Boelens WC, Leunissen JA, de Jong WW (2003) The human genome encodes 10 alpha-crystallin-related small heat shock proteins: HspB1-10. Cell Stress Chaperones 8(1):53–61

    Article  PubMed  CAS  Google Scholar 

  • Kappe G, Boelens WC, de Jong WW (2010) Why proteins without an alpha-crystallin domain should not be included in the human small heat shock protein family HSPB. Cell Stress Chaperones 15(4):457–461

    Article  PubMed  CAS  Google Scholar 

  • Kasakov AS, Bukach OV, Seit-Nebi AS, Marston SB, Gusev NB (2007) Effect of mutations in the beta5–beta7 loop on the structure and properties of human small heat shock protein HSP22 (HspB8, H11). FEBS J 274(21):5628–5642

    Article  PubMed  CAS  Google Scholar 

  • Kato K, Shinohara H, Goto S, Inaguma Y, Morishita R, Asano T (1992) Copurification of small heat shock protein with alpha B crystallin from human skeletal muscle. J Biol Chem 267(11):7718–7725

    PubMed  CAS  Google Scholar 

  • Kato K, Goto S, Inaguma Y, Hasegawa K, Morishita R, Asano T (1994) Purification and characterization of a 20-kDa protein that is highly homologous to alpha B crystallin. J Biol Chem 269(21):15302–15309

    PubMed  CAS  Google Scholar 

  • Kazakov AS, Markov DI, Gusev NB, Levitsky DI (2009) Thermally induced structural changes of intrinsically disordered small heat shock protein Hsp22. Biophys Chem 145(2–3):79–85

    Article  PubMed  CAS  Google Scholar 

  • Kim KK, Kim R, Kim SH (1998) Crystal structure of a small heat-shock protein. Nature 394(6693):595–599

    Article  PubMed  CAS  Google Scholar 

  • Kriehuber T, Rattei T, Weinmaier T, Bepperling A, Haslbeck M, Buchner J (2010) Independent evolution of the core domain and its flanking sequences in small heat shock proteins. FASEB J 24(10):3633–3642

    Article  PubMed  CAS  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227(5259):680–685

    Article  PubMed  CAS  Google Scholar 

  • Laganowsky A, Benesch JL, Landau M, Ding L, Sawaya MR, Cascio D, Huang Q, Robinson CV, Horwitz J, Eisenberg D (2010) Crystal structures of truncated alphaA and alphaB crystallins reveal structural mechanisms of polydispersity important for eye lens function. Protein Sci 19(5):1031–1043

    Article  PubMed  CAS  Google Scholar 

  • Lelj-Garolla B, Mauk AG (2006) Self-association and chaperone activity of Hsp27 are thermally activated. J Biol Chem 281(12):8169–8174

    Article  PubMed  CAS  Google Scholar 

  • Louapre P, Grongnet JF, Tanguay RM, David JC (2005) Effects of hypoxia on stress proteins in the piglet heart at birth. Cell Stress Chaperones 10(1):17–23

    Article  PubMed  CAS  Google Scholar 

  • Lutsch G, Vetter R, Offhauss U, Wieske M, Grone HJ, Klemenz R, Schimke I, Stahl J, Benndorf R (1997) Abundance and location of the small heat shock proteins HSP25 and alphaB-crystallin in rat and human heart. Circulation 96(10):3466–3476

    PubMed  CAS  Google Scholar 

  • McHaourab HS, Godar JA, Stewart PL (2009) Structure and mechanism of protein stability sensors: chaperone activity of small heat shock proteins. Biochemistry 48(18):3828–3837

    Article  PubMed  CAS  Google Scholar 

  • Mymrikov EV, Bukach OV, Seit-Nebi AS, Gusev NB (2010) The pivotal role of the beta 7 strand in the intersubunit contacts of different human small heat shock proteins. Cell Stress Chaperones 15(4):365–377

    Article  PubMed  CAS  Google Scholar 

  • Narberhaus F (2002) Alpha-crystallin-type heat shock proteins: socializing minichaperones in the context of a multichaperone network. Microbiol Mol Biol Rev 66(1):64–93

    Article  PubMed  CAS  Google Scholar 

  • Rogalla T, Ehrnsperger M, Preville X, Kotlyarov A, Lutsch G, Ducasse C, Paul C, Wieske M, Arrigo AP, Buchner J, Gaestel M (1999) Regulation of Hsp27 oligomerization, chaperone function, and protective activity against oxidative stress/tumor necrosis factor alpha by phosphorylation. J Biol Chem 274(27):18947–18956

    Article  PubMed  CAS  Google Scholar 

  • Sakuma K, Watanabe K, Totsuka T, Kato K (1998) Pathological changes in levels of three small stress proteins, alphaB crystallin, HSP 27 and p20, in the hindlimb muscles of dy mouse. Biochim Biophys Acta 1406(2):162–168

    PubMed  CAS  Google Scholar 

  • Schuck P (2000) Size-distribution analysis of macromolecules by sedimentation velocity ultracentrifugation and lamm equation modeling. Biophys J 78(3):1606–1619

    Article  PubMed  CAS  Google Scholar 

  • Shemetov AA, Gusev NB (2011) Biochemical characterization of small heat shock protein HspB8 (Hsp22)-Bag3 interaction. Arch Biochem Biophys 513(1):1–9

    PubMed  CAS  Google Scholar 

  • Sobott F, Benesch JL, Vierling E, Robinson CV (2002) Subunit exchange of multimeric protein complexes. Real-time monitoring of subunit exchange between small heat shock proteins by using electrospray mass spectrometry. J Biol Chem 277(41):38921–38929

    Article  PubMed  CAS  Google Scholar 

  • Stamler R, Kappe G, Boelens W, Slingsby C (2005) Wrapping the alpha-crystallin domain fold in a chaperone assembly. J Mol Biol 353(1):68–79

    Article  PubMed  CAS  Google Scholar 

  • Studer S, Narberhaus F (2000) Chaperone activity and homo- and hetero-oligomer formation of bacterial small heat shock proteins. J Biol Chem 275(47):37212–37218

    Article  PubMed  CAS  Google Scholar 

  • Sugiyama Y, Suzuki A, Kishikawa M, Akutsu R, Hirose T, Waye MM, Tsui SK, Yoshida S, Ohno S (2000) Muscle develops a specific form of small heat shock protein complex composed of MKBP/HSPB2 and HSPB3 during myogenic differentiation. J Biol Chem 275(2):1095–1104

    Article  PubMed  CAS  Google Scholar 

  • Sun X, Fontaine JM, Rest JS, Shelden EA, Welsh MJ, Benndorf R (2004) Interaction of human HSP22 (HSPB8) with other small heat shock proteins. J Biol Chem 279(4):2394–2402

    Article  PubMed  CAS  Google Scholar 

  • Suzuki A, Sugiyama Y, Hayashi Y, Nyu-i N, Yoshida M, Nonaka I, Ishiura S, Arahata K, Ohno S (1998) MKBP, a novel member of the small heat shock protein family, binds and activates the myotonic dystrophy protein kinase. J Cell Biol 140(5):1113–1124

    Article  PubMed  CAS  Google Scholar 

  • Taylor RP, Benjamin IJ (2005) Small heat shock proteins: a new classification scheme in mammals. J Mol Cell Cardiol 38(3):433–444

    Article  PubMed  CAS  Google Scholar 

  • Treweek TM, Rekas A, Lindner RA, Walker MJ, Aquilina JA, Robinson CV, Horwitz J, Perng MD, Quinlan RA, Carver JA (2005) R120G alphaB-crystallin promotes the unfolding of reduced alpha-lactalbumin and is inherently unstable. FEBS J 272(3):711–724

    Article  PubMed  CAS  Google Scholar 

  • van Montfort RL, Basha E, Friedrich KL, Slingsby C, Vierling E (2001) Crystal structure and assembly of a eukaryotic small heat shock protein. Nature Struct Biol 8(12):1025–1030

    Article  PubMed  Google Scholar 

  • Verschuure P, Tatard C, Boelens WC, Grongnet JF, David JC (2003) Expression of small heat shock proteins HspB2, HspB8, Hsp20 and cvHsp in different tissues of the perinatal developing pig. Eur J Cell Biol 82(10):523–530

    Article  PubMed  CAS  Google Scholar 

  • Vos MJ, Hageman J, Carra S, Kampinga HH (2008) Structural and functional diversities between members of the human HSPB, HSPH, HSPA, and DNAJ chaperone families. Biochemistry 47(27):7001–7011

    Article  PubMed  CAS  Google Scholar 

  • Vos MJ, Zijlstra MP, Carra S, Sibon OC, Kampinga HH (2011) Small heat shock proteins, protein degradation and protein aggregation diseases. Autophagy 7(1):101–103

    Article  PubMed  Google Scholar 

  • Zantema A, Verlaan-De Vries M, Maasdam D, Bol S, van der Eb A (1992) Heat shock protein 27 and alpha B-crystallin can form a complex, which dissociates by heat shock. J Biol Chem 267(18):12936–12941

    PubMed  CAS  Google Scholar 

  • Zavialov A, Benndorf R, Ehrnsperger M, Zav’yalov V, Dudich I, Buchner J, Gaestel M (1998) The effect of the intersubunit disulfide bond on the structural and functional properties of the small heat shock protein Hsp25. Int J Biol Macromol 22(3–4):163–173

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

This investigation was supported by Russian Foundation for Basic Science (grant 10-04-00026).

Author information

Authors and Affiliations

  1. Department of Biochemistry, School of Biology, Moscow State University, Moscow, 119991, Russian Federation

    Evgeny V. Mymrikov, Alim S. Seit-Nebi & Nikolai B. Gusev

Authors
  1. Evgeny V. Mymrikov
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Alim S. Seit-Nebi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Nikolai B. Gusev
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Nikolai B. Gusev.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Online Resource 1

Reduction of disulfide cross-linked wild-type HspB1 and Cys mutants of human small heat shock proteins. Isolated small heat shock proteins (lanes 1 and 3) or their pair-wise mixture (lane 2) were subjected to subunit exchange followed by mild oxidation as described in “Material and methods”. The samples thus obtained were subjected to reduction and SDS–gel electrophoresis. a HspB1 + HspB6; b HspB5 + HspB6; c HspB1 + HspB5; d HspB8 + HspB5; e HspB8 + HspB6; f HspB8 + HspB1. Trace amounts of non-reduced disulfide cross-linked dimers are detected as a very faint bands with apparent molecular masses of 42–66 kDa. The positions of molecular mass standards (in kilodaltons) are indicated by arrows (PDF 233 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Mymrikov, E.V., Seit-Nebi, A.S. & Gusev, N.B. Heterooligomeric complexes of human small heat shock proteins. Cell Stress and Chaperones 17, 157–169 (2012). https://doi.org/10.1007/s12192-011-0296-0

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12192-011-0296-0

Keywords

Search

Navigation