The MitCHAP-60 disease is due to entropic destabilization of the human mitochondrial Hsp60 oligomer.
The 60-kDa heat shock protein (mHsp60) is a vital cellular complex that mediates the folding of many of the mitochondrial proteins. Its function is executed in cooperation with the co-chaperonin, mHsp10, and requires ATP. Recently, the discovery of a new mHsp60-associated neurodegenerative disorder, MitCHAP-60 disease, has been reported. The disease is caused by a point mutation at position 3 (D3G) of the mature mitochondrial Hsp60 protein, which renders it unable to complement the deletion of the homologous bacterial protein in Escherichia coli (Magen, D., Georgopoulos, C., Bross, P., Ang, D., Segev, Y., Goldsher, D., Nemirovski, A., Shahar, E., Ravid, S., Luder, A., Heno, B., Gershoni-Baruch, R., Skorecki, K., and Mandel, H. (2008) Am. J. Hum. Genet. 83, 30-42). The molecular basis of the MitCHAP-60 disease is still unknown. In this study, we present an in vitro structural and functional analysis of the purified wild-type human mHsp60 and the MitCHAP-60 mutant. We show that the D3G mutation leads to destabilization of the mHsp60 oligomer and causes its disassembly at low protein concentrations. We also show that the mutant protein has impaired protein folding and ATPase activities. An additional mutant that lacks the first three amino acids (N-del), including Asp-3, is similarly impaired in refolding activity. Surprisingly, however, this mutant exhibits profound stabilization of its oligomeric structure. These results suggest that the D3G mutation leads to entropic destabilization of the mHsp60 oligomer, which severely impairs its chaperone function, thereby causing the disease.
Parnas
Avital
A
Department of Biochemistry, George S. Wise Faculty of Life Sciences, Tel Aviv University, 69778 Tel Aviv.
Nadler
Michal
M
Department of Structural Biology, Weizmann Institute of Science, Rehovot 76100.
Nisemblat
Shahar
S
Department of Biochemistry, George S. Wise Faculty of Life Sciences, Tel Aviv University, 69778 Tel Aviv.
Horovitz
Amnon
A
Department of Structural Biology, Weizmann Institute of Science, Rehovot 76100. Electronic address: amnon.horovitz@weizmann.ac.il.
Mandel
Hanna
H
Rappaport Faculty of Medicine and Research Institute, Technion-Israel Institute of Technology and Metabolic Disease Unit, Rambam Health Care Campus, Haifa 31096, Israel.
Azem
Abdussalam
A
Department of Biochemistry, George S. Wise Faculty of Life Sciences, Tel Aviv University, 69778 Tel Aviv. Electronic address: azema@tauex.tau.ac.il.
D000595:Amino Acid Sequence; D000818:Animals; D018834:Chaperonin 60 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019277:Entropy; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D019636:Neurodegenerative Diseases / Q000378:metabolism*; D017354:Point Mutation; D017510:Protein Folding; D020836:Protein Structure, Quaternary
D018834:Chaperonin 60; D024101:Mitochondrial Proteins
Curr Opin Struct Biol. 1998 Feb;8(1):93-100
9519301
J Biol Chem. 2007 Feb 16;282(7):4463-4469
17178727
Curr Opin Struct Biol. 2005 Dec;15(6):646-51
16249079
Am J Hum Genet. 2008 Jul;83(1):30-42
18571143
EMBO J. 1999 Apr 15;18(8):2049-56
10205159
Nature. 1989 Sep 14;341(6238):125-30
2528694
J Biol Chem. 2004 Nov 12;279(46):47906-11
15371451
J Biol Chem. 1989 Jul 15;264(20):12001-8
2568357
J Neurol. 2007 Jul;254(7):897-900
17420924
J Biol Chem. 1994 Oct 28;269(43):26858-64
7929423
J Biol Chem. 2001 Feb 16;276(7):4981-7
11050098
J Biol Chem. 1993 May 15;268(14):9957-9
8098040
J Biol Chem. 1995 Sep 1;270(35):20404-9
7657615
Biochemistry. 1998 Jul 21;37(29):10370-80
9671505
Proc Natl Acad Sci U S A. 1997 Apr 15;94(8):3576-8
9108018
FEBS Lett. 2002 Oct 2;529(1):1-5
12354603
Nature. 1993 Jul 15;364(6434):255-8
8100614
Methods Enzymol. 1998;290:203-17
9534164
Crit Rev Biochem Mol Biol. 2006 Jul-Aug;41(4):211-39
16849107
J Biol Chem. 1986 Sep 15;261(26):12414-9
3017973
J Biol Chem. 2008 Jun 6;283(23):15694-700
18400758
Annu Rev Cell Dev Biol. 2007;23:115-45
17489689
Proc Natl Acad Sci U S A. 1996 Oct 15;93(21):11622-7
8876186
Nature. 1996 Jun 13;381(6583):571-9
8637592
Biochemistry. 1994 Jul 12;33(27):8262-71
8031761
Mol Cell. 1998 Jul;2(1):93-9
9702195
Eur J Biochem. 2001 Jun;268(12):3465-72
11422376
Am J Hum Genet. 2002 May;70(5):1328-32
11898127
Cardiovasc Toxicol. 2003;3(3):263-8
14555791
Nature. 1989 Feb 16;337(6208):620-5
2645524
Cancer Biol Ther. 2007 Apr;6(4):487-9
17457039
Chem Rev. 2006 May;106(5):1917-30
16683761
Proc Natl Acad Sci U S A. 1993 Dec 1;90(23):10967-71
7902576
Biochemistry. 1995 Apr 25;34(16):5303-8
7727391
Cell Stress Chaperones. 2009 Sep;14(5):509-19
19224397
J Mol Biol. 1993 May 5;231(1):58-64
8098773
Cell Stress Chaperones. 2006 Summer;11(2):116-28
16817317
Protein Sci. 2007 Feb;16(2):316-22
17242434
Proc Natl Acad Sci U S A. 1997 Apr 15;94(8):3571-5
9108017