A Multipronged Method for Unveiling Subtle Structural–Functional Defects of Mutant Chaperone Molecules Causing Human Chaperonopathies
Chaperonopathies are diseases in which abnormal chaperones play an etiopathogenic role. A chaperone is mutated or otherwise abnormal (e.g., modified by an aberrant posttranslational modification) in structure/function. To understand the pathogenic mechanisms of chaperonopathies, it is necessary to elucidate the impact of the pathogenic mutation or posttranslational modification on the chaperone molecule’s properties and functions. This impact is usually subtle because if it were more than subtle the overall effect on the cell and organism would be catastrophic, lethal. This is because most chaperones are essential for life and, if damaged in structure/function too strongly, there would be death of the cell/organism, and no phenotype, i.e., there would be no patients with chaperonopathies. Consequently, diagnostic procedures and analysis of defects of the abnormal chaperones require a multipronged method for assessing the chaperone molecule from various angles. Here, we present such a method that includes assessing the intrinsic properties and the chaperoning functions of chaperone molecules.
KeywordsChaperonopathies Genetic chaperonopathies Mutant chaperones Subtle changes Multipronged method Molecular properties Chaperoning functions Calorimetry Heat protection Fibril dispersion
Part of the work by PLSB and DB was carried out using instruments funded by PO. FESR 2007/2013 “Piattaforma regionale di ricerca traslazionale per la salute” 4.1.2. Asse IV. We thank Drs. A. Provenzano and F. Impallari for technical support.
Part of this work was carried out at the Biomolecular Labeling Laboratory-National Institute of Standards and Technology (NIST), Rockville, USA. FTR, ECdeM, and AJLM thank Zvi Kelman for his hospitality and stimulating discussions.
FTR was supported by NSF USA grants EAR0747394, EAR0747412, and MCB0605301; and Air Force Office of Scientific Research under Grants AFOSR 03-S-28900; and 496 9550-10-1-0272 (IKL and FTR). IKL was supported in part by the National Science Foundation under Grant No. CHE-1152752 (I.K.L.). AJLM and ECdeM were partially supported by IMET. This work was done under the agreement between IEMEST (Italy) and IMET (USA) (IMET contribution number IMET 18-003).
- 2.Macario AJL, Conway de Macario E (2016) The chaperoning and the immune systems with the microbiome integrate a matrix that supports health: when one of them is disturbed the others suffer and disease ensues. Life Safety Security (LiSS) 4:101–123 http://www.iemest.eu/life-safety-and-security/images/Doc/ARTICOLI/2016/macario_24/Macario04.pdfGoogle Scholar
- 5.Conway de Macario E, Robb FT, Macario AJL (2017) Prokaryotic chaperonins as experimental models for elucidating structure-function abnormalities of human pathogenic mutant counterparts. Front Mol Biosci 3:84. https://doi.org/10.3389/fmolb.2016.00084 https://www.ncbi.nlm.nih.gov/pubmed/28119916 http://journal.frontiersin.org/article/10.3389/fmolb.2016.00084/full CrossRefPubMedPubMedCentralGoogle Scholar
- 7.Sturtevant JM (1987) Biochemical applications of differential scanning calorimetry. Ann Rev Phys Chem 38:463–488. https://doi.org/10.1146/annurev.pc.38.100187.002335 CrossRefGoogle Scholar
- 8.Freire E (1995) Thermal denaturation methods in the study of protein folding. In: Abelson J, Simon M, Johnson M, Ackers G (eds) Methods in enzymology, Energetics of biological macromolecules, vol 259. Academic Press, San Diego, pp 144–168Google Scholar
- 11.Velazquez-Campoy A, Leavitt SA, Freire E (2015) Characterization of protein-protein interactions by isothermal titration calorimetry. In: Meyerkord CL, Fu H (eds) Methods in molecular biology, Protein-protein interactions: methods and applications, vol 1278, 2nd edn. Springer, New York, pp 183–204Google Scholar
- 13.Min W, Angileri F, Luo H, Lauria A, Shanmugasundaram M, Almerico AM, Cappello F, Conway de Macario E, Lednev IK, Macario AJL, Robb FT (2014) A human CCT5 gene mutation causing distal neuropathy impairs hexadecamer assembly in an archaeal model. Sci Rep 4:6688. https://doi.org/10.1038/srep06688 CrossRefPubMedPubMedCentralGoogle Scholar
- 16.Conway KA, Lee SJ, Rochet JC, Ding TT, Williamson RE, Lansbury PT (2000) Acceleration of oligomerization, not fibrillization, is a shared property of both α-synuclein mutations linked to early-onset Parkinson’s disease: Implications for pathogenesis and therapy. Proc Natl Acad Sci U S A 97(2):571–576CrossRefGoogle Scholar
- 24.Spigolon D, Gallagher DT, Velazquez-Campoy A, Bulone D, Narang J, San Biagio PL, Cappello F, Macario AJL, Conway de Macario E, Robb FT (2017) Quantitative analysis of the impact of a human pathogenic mutation on the CCT5 chaperonin subunit using a proxy archaeal ortholog. Biochem Biophys Rep 12:66–71, 2017. https://doi.org/10.1016/j.bbrep.2017.07.011 CrossRefPubMedPubMedCentralGoogle Scholar