Regulation of Muscle Contraction and How Mutations in the Muscle Proteins Cause Heart Disease: The Laboratory Techniques Involves Molecular Biology, Protein Expression, Purification and Characterization Using Biochemical and Biophyisical Techniques
DOI:
https://doi.org/10.14738/bjhmr.116.14733Keywords:
hypertrophic cardiomyopathy, Tropomyosin, Troponin, Myosin, mutations, autosomal-dorminant, inheritance, autosomal-recessive, Sarcomere, Arrhythmogenic, Proteins, etcAbstract
The expansion of the causative mutations to the rigid thin filament changed the description of hypertrophic cardiomyopathy (HCM) from an illness of the cardiac motor to a syndrome of the cardiac sarcomere and significantly extended the gasp of the potential molecular pathogenic mechanism. An interesting hypotheses concerning disease mechanism posted that the diverse medical prognoses in the familial type of HCM may possibly be related to mutations in self-regulating protein machinery of the sarcomere (Frank et al, 1968). The journal of the novel study in 1990 establishes the inherited association of the beta-myosin and tropomyosin heavy chain genetic material to familial type of hypertrophic cardiomyopathy (HCM). The current studies conducted by some researchers elaborated on the various genetic alterations inside the genes encoding for the sarcomeric cardiac proteins, alpha tropomyosin, troponin T, and myosin protein components. The regularity of gene alteration in the alpha tropomyosin protein (TPM1) is lesser, contributing to 5% of FHC. Currently, the D175N gene mutation has been recognized in various unrelated populations, signifying that this spot could be an abnormal gene “hot spot” for the disease. In this project research, a wild type of normal protein and mutant genetic proteins (E180G and D175N) which are clinically involved in familial hyper cardiomyopathy (FHCM) were produced. Having in mind that the main effect of mutations E180G and D175N are mainly related to the thermal stability of the protein; this research will also investigate the differences between the thermal stability of wild type and mutated protein types using a Dye base fluorescent method of analysis. Dye based fluorescent method was used to monitor protein folding as a function of temperature for wild type tropomyosin and for HCM mutant E180G and D175N proteins. The column chromatography method of purification was used to purify the wild type and mutated proteins, and the protein bands were separated using gel electrophoresis methods. A similar assessment of folding stability and structural reports of several authors was in consistency with this present report which suggested that such mutations might alter protein folding. The results agree with previously published reports on the impaired function of expressed E180G and D175N mutations suggesting that the biochemical defects of the motor domain may affect myosin filament assembly in the sarcomere. For future prospects, future biochemical analysis of several other FHC mutations will be needed to establish a definite correlation between the enzymatic impairment between different mutants and their clinical phenotype of heart disease.
References
Aminetzach, Yael T.; Macpherson, J. Michael; Petrov, Dmitri A. (July 29, 2005). "Pesticide Resistance via Transposition-Mediated Adaptive Gene Truncation in Drosophila".
Washington, D.C.: American Association for the Advancement of Science) 309 (5735): 764– 767. Boillée, Séverine; Vande Velde, Christine; Cleveland, Don W. (October 5, 2006). "ALS: A Disease of Motor Neurons and Their Nonneuronal Neighbors”. (Cambridge, MA: Cell Press) 52 (1): 39–59.
Boillée, Séverine; Vande Velde, Christine; Cleveland, Don W. (October 5, 2006). "ALS: A Disease of Motor Neurons and Their Nonneuronal Neighbors". Neuron (Cambridge, MA: Cell Press) 52 (1): 39–59. doi: 10.1016/j.neuron.2006.09.018. ISSN 0896-6273. PMID 17015226.
Bertram, John S. (December 2000). "The molecular biology of cancer". Molecular Aspects of Medicine (Amsterdam, the Netherlands: Elsevier) 21 (6): 167–223.
Bing W, Redwood CS, Purcell IF, et al. Effects of two hypertrophic cardiomyopathy mutations in alphatropomyosin, Asp175Asn and Glu180Gly, on Ca21 regulation of thin filament motility. Biochem Biophys Res Commun. 1997; 236:760–764.
Charron P, Dubourg O, Desnos M, et al. Clinical features and prognostic implications of familial hypertrophic cardiomyopathy related to the cardiac-myosin-binding protein C gene. Circulation. 1998;97: 2230–2236.
Circulation. 1998;97: 2230–2236.
Charron P, Dubourg O, Desnos M, et al. Clinical features and prognostic implications of familial hypertrophic cardiomyopathy related to the cardiac-myosin-binding protein C gene. Doolittle and Yanofsky, (1968). Mutants of Escherichia coli with an altered tryptophanyl- transfer ribonucleic acid synthetase. Journal of bacteriology (impact factor: 2.81): 95(4)1283- 94.
Davies, M. J. 1984. The current status of myocardial disarray in hypertrophic cardiomyopathy. Br. Heart. J.
Dausse, E., and K. Schwartz. 1993. Genetic heterogeneity of familial hypertrophic cardiomyopathy. Neuromuscul. Disord. 3:483–486.
Ebashi S, Kodama A, Ebashi F, (1968). Troponin 1. Preparation and physiological function. J Biochem. 64:465-
Elvin CM, Thompson PR, Argall ME, Hendry P, Stamford NP, Lilley P, Dixon NE, (1990). Modified bacteriophage lambda promoter vectors for overproduction of proteins in E.Coli. Research schho of chemistry, Australian National university, Canberra. 1: 87(1):123-6.
Eldin P, Le Cunff M, Diederich KW, Jaenicke T, Cornillon B, Mornet D, Vosberg HP, Leger JJ, (1990). Expression of human beta-myosin heavy chain fragments in E.Coli: localization of actin interfaces on cardiac myosin. J Muscle Res Cell Motil. 11:378-391.
Freese, Ernst (April 15, 1959). "The difference between Spontaneous and Base-Analogue Induced Mutations of Phage T4". Proc. Natl. Acad. Sci. U.S.A. (Washington, D.C.: National Academy of Sciences) 45 (4): 622–633. (1): 1–5.
Maluze, U. O. (2024). Regulation of Muscle Contraction and How Mutations in the Muscle Proteins Cause Heart Disease. The Laboratory Techniques Fananapazir L, Epstein ND. Genotype-phenotype correlations in hypertrophic cardiomyopathy: insights provided by comparisons of kindreds with distinct and identical b - myosin heavy chain gene mutations. Circulation. 1994; Freese, Ernst (June 1959). "The specific mutagenic effect of base analogues on Phage T4". Journal of Molecular Biology (Amsterdam, the Netherlands: Elsevier) 1 (2): 87–105.
Frank S, Braunwald E. (1968). Idiopathic subaortic stenosis. Clinical analysis of 126 patients with emphasis on the natural history. Circulation. 37:759-788.
Golitsina N, An Y, Greenfield NJ, et al. Effects of two familial hypertrophic cardiomyopathy- causing mutations on alpha-tropomyosin structure and function [published erratum appears in Biochemistry. 1999; 38:3850].
Biochemistry. 1997; 36:4637–4642.
Glascock and Weickert, (1998). Improving the Lac system for synthetic biology.
Geisterfer-Lowrance AA, Kass S, Tanigawa G, Vosberg HP, McKenna W, Seidman CE, Seidman JG, (1990). A molecular basis for familial hypertrophic cardiomyopathy: a better cardiac muosin heavy chain gene missence mutation. Cell; 62:999-1006.
Hitchcock SE, Huxley HE, Szent-Gyorgyi AG, (1973). Calcium sensitive binding of troponin to actin-tropomyosin: a two-site model for troponin action. J Mol Biol. 80:825-836.
Hogan, C. Michael (October 12, 2010). "Mutation". In Monosson, Emily. Encyclopedia of Earth. Washington, D.C.: Environmental Information Coalition, National Council for Science and the Environment. Retrieved 2015-10-08. Imura A, Harada H, Park JE, Nishi H, Satoh M, Takahashi M, Hirobi S, Sasaoka T, Ohbuchi N, Nakamura T, Koyanagi T, Hwang TH, Choo JA, Chung KS, Hasegawa A, Nagai R, Okaziki O, Nakamura H, Matsuzaki M, Sakamoto
T, Toshima H, Koga Y, Imaizumi T, Sasazuki T. (1997). Mutations in the cardiac troponin I gene associated with hypertrophic cardiomyopathy. Nature, 16:379-382.
Jongbloed RJ; Marcelis, CL; Doevendans A.P; Schmeitz-Mulkens, J.M; Van Dockum, W.G. & Geraedts, J.P. (2003). Variable clinical manifestation of a novel missense mutation in the alpha tropomyosin (TPM1) gene in familial hypertrophic cardiomyopathy. J.Am. Coll. Cardiol. 41, 981-986.
Kannon T, Chang S, Seidman JG, Seidman CE, (2010). Genetics of hypertrophic cardiomyopathy. Curr Opin Cardiol. PMID 20124998; PMCID 2932754.
Kamisago M, Sharma SD, DePalma SR, Solomon S, Sharma P, McDonough B, Smoot L, Mullen MP, Woolf PK, Wigle ED, Seidman JG, Seidman CE. Mutations in sarcomere protein genes as a cause of dilated cardiomyopathy. N Engl J Med. 343:1688-1696.
Kunkel, T.A. (1985) Proc Natl Acad Sci USA 82(2):488-92.
Kilbey, B. J. (1995). "Charlotte Auerbach)". Genetics 141
Landstrom AP, Parvatiyar MS, Pinto JR, Marquardt ML, Bos JM, Tester DJ, Ommen SR, Potter JD, Ackerman MJ. (2008). Molecular and functional characterization of novel hypertrophic cardiomyopathy susceptibility mutations in TNNC1-encoded troponin C. J Mol Cell Cardiol. 45:281-288.
Lees-Miller JP, Helfman DM. The molecular basis for tropomyosin isoform diversity. Bio Essays. 1991; 13:429–
McKillop DF, Geeves MA, (1993). Regulation of the interaction between actin and myosin subfragment I: evidence for thre states of the thin filament. Byophys J. 65:693-701. 185
Mogensen J, Murphy RT, Shaw T, Bahi A, Redwood C, Watkins H, Burke M, Elliott PM, McKenna WJ. (2004). Severe disease expression of cardiac troponin C and T mutations in patients with idiopathic dilated cardiomyopathy. J Am Coll Cardiol; 44:2033-2040.
Maron, B. J., R. O. Bonow, R. O. Cannon, M. B. Leon, and S. E. Epstein. 1987. Hypertrophic cardiomyopathy: interrelations of clinical manifestations, pathophysiology and therapy. N. Engl. J. Med 316:780–789.
Maron, B. J., W. C. Roberts, and S. E. Epstein. 1986. Causes of sudden death in competitive athletes. J. Am. Coll. Cardiol. 7:204–214.
Müller-Hill, B.L. Crao, W. Gilbert (1968). Mutant that makes more lac repressor.Proc.Natl.Acad.Sci.USA 59:1259-Nakajima-Taniguchi C, Matsui H, et al. Novel missense mutation in alpha tropomyosin gene found in Japanese patients with hypertrophic cardiomyopathy. J Moll Cell Cardiol. 1995; 27:2053–2058.
Nelson M; McClelland M. (1992) Methods Enzymol 216:279-303.
Olson TM, Doan TP, Doan TP, Kishimoto NY, Whitby FG, Ackerman MJ, Fananapazir L. (2000). Inherited and de novo mutations in the cardiac actin gene cause hypertrophic cardiomyopathy. J Mol Cell Cardiol. 32:1678-1694.
Preston LC, Lipscomb S, Robinson P, Mogensen J, McKenna WJ, Watkins H, Ashley CC, Redwood CS, (2007). Functional effects of the DCM mutant Gly159Asp troponin C in skinned muscle fibres. Pflugers Arch. 453:771-
Papworth C; Bauer J.C; Braman J; Wright D.A. (1996) strategies 9(3):3-4.
Rose and Yanofsky, (1974). Interaction of the operator of the tryptophan operon with repressor. Proc. Natl. Acad.Sci. 713134-3138.
Regitz-Zagrosek, V; Erdmann, J; Wellnhofer, E; Raibe, J. & Fleck, E. (2000). Novel mutation in the alpha tropomyosin gene and transition from hypertrophic dilated cardiomyopathy. Circulation, 102, 112-116. Reed PW, Davies JL, Copeman JB, et al. Chromosome-specific microsatellite sets for fluorescent-based, semiautomated genome mapping. Nat Genet. 1994; 7:390–395.
Solomon, S. D., A. A. T. Geisterfer-Lowrance, H. P. Vosberg, G. Hiller, J. A. Jarcho, C. C. Morton, W. O. McBride, A. L. Mitchell, A. E. Bale, W. J. McKenna, J. G. Seidman, and C. E. Seidman. 1990. A locus for familial hypertrophic cardiomyopathy is closely linked to the cardiac myosin heavy chain genes, CRI-L436, and CRI-L329 on chromosome 14 at q11–q12. Am. J. Hum. Genet. 47:389–394.
Shortle, D.; Dimaio, D.; Nathans, D. (1981). "Directed Mutagenesis". Annual Review of Genetics 15: 265–294.
Sweeny HL, Straceski AJ, Leinwand LA, Tikunov BA, Faust L, (1994). Heterologous expression of a cardiomyopathic myosin that is defective in its actin interaction. J Biol Chem. 269:1603-1605.
Schwartz K, Carrier L, Guicheney P, et al. Molecular basis of familial cardiomyopathies. Circulation. 1995; Satoh M, Takahashi, Sakamoto T, et al. Structural analysis of the titin gene in hypertrophic cardiomyopathy: identification of a novel disease gene. Biochem Biophys Res Commun. 1999; 262:411–417.
Thierfelder, L., H. Watkins, C. MacRae, R. Lamas, W. McKenna, H. P. Vosberg, J. G. Seidman, and C. E. Seidman. 1994. Alpha-tropomyosin and cardiac troponin T mutations cause familial hypertrophic cardiomyopathy: a disease of the sarcomere. Cell. 77:701–712. 186
Maluze, U. O. (2024). Regulation of Muscle Contraction and How Mutations in the Muscle Proteins Cause Heart Disease. The Laboratory Techniques
Taylor, J.W; Ott, J; Eckstein F. (1985) Nucleic Acids Res 13 (24):8765-85
Teare D. (1958). Asymmetrical hypertrophy of the heart in young adults. Br Heart J. 1958; 20:1-8
U Q, Dewey S, Nguyen S, Gomes AV, (2010). Malignant and benign mutations in familial cardiomyopathies: insights into mutations linked to complex cardiovascular phenotypes. J Mol Cell Cardiol; 48:899-909.
Van Driest, S.L; Will, M.L; Atkins, D.L. & Ackerman, M.J. (2002). A novel TPMI mutation in a family with hypertrophic cardiomyopathy and sudden cardiac death in childhood. Am.J. Cardiol. 90, 1123-1127.
Vandeyar, M.A; Weiner M.p; Hutton C.J; Batt, C.A. (1988) Gene 65(1):129-33.
Watkins H, Anan R, Coviello DA, et al. A de novo mutation in a -tropomyosin that causes hypertrophic cardiomyopathy. Circulation. 1995; 91:2302–2305.
Watkins, H., W. J. McKenna, L. Thierfelder, H. J. Suk, R. Anan, A. O’Donoghue, P. Spirito, A. Matsumori, C. S. Moravec, J. G. Seidman, and C. E. Seidman. 1995b. Mutations in the genes for cardiac troponin T and alpha tropomyosin in hypertrophic cardiomyopathy. N. Engl. J. Med. 332:1058–1064.
Zot AS, Potter JD. Structural aspects of troponin-tropomyosin regulation of skeletal muscle contraction. Annu Rev Biophys Chem. 1987;16: 535–559. 10. Tobacman LS. Thin filament- mediated regulation of cardiac contraction.
Annu Rev Physiol. 1996; 58:447–481.
Downloads
Published
How to Cite
Issue
Section
License
Copyright (c) 2024 Ugochukwu Obinna Maluze
This work is licensed under a Creative Commons Attribution 4.0 International License.
