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Effects of Myopathy-causing Mutations on Tropomyosin Structure and Function

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posted on 04.09.2017, 10:33 authored by Saeed Ahmed Asiri
Tropomyosin determinants for actin binding have not been identified completely and the nature and position of residues involved in thin filament dynamics has not been established. To date a number of Tropomyosin mutations have been linked to several muscle diseases including cardiomyopathies and skeletal muscle myopathies. In this thesis, we aimed to investigate the following tropomyosin mutations R90G, E163K, R167G, E240K, R244G and M2811 which have been shown to cause several severe skeletal muscle myopathies. We used various structural, biochemical and kinetic methods to assess the impact of these mutations on tropomyosin structure and biochemical properties. Fluorescence emission spectroscopy, and transient kinetics were used to assess the effect of these mutations on the equilibrium distribution and kinetics of transitions between different thin filament regulatory states. Overall the data demonstrated that: 1) all tropomyosin mutations except (E163K, E240K, M2811 and R90GR167G) affected the thermal stability of tropomyosin but not the a-helical coiled coil structure. 2) The size of the cooperative unit n was reduced by all tropomyosin mutations. 3) Tropomyosin mutations did not affect the proportion of thin filaments in the blocked state (at low Ca2+). 4) Tropomyosin mutations did affect the maximum observed rate constant of thin filament transition between the ON and OFF states. 5) Several tropomyosin mutations have affected tropomyosin-troponin binding affinity but none of the mutations had any effect on the actin binding affinity. Overall these results provide insight into the mechanism by which tropomyosin bind actin and troponin, tropomyosin related thin filament cooperativity and allosteric transitions.



El-Mezgueldi, Mohammed

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Department of Molecular and Cell Biology

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University of Leicester

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