The effects of cardiomyopathy-associated mutations in the head-to-tail overlap junction of α-tropomyosin on its properties and interaction with actin

Matyushenko, Alexander M.; Koubassova, Natalia A.; Shchepkin, Daniil V.; Kopylova, Galina V.; Nabiev, Salavat R.; Никитина, Л. В.; Bershitsky, Sergey Y.; Levitsky, Dmitrii I.; Tsaturyan, Andrey K.

doi:10.1016/j.ijbiomac.2018.09.105

Cited by 21 publications

(38 citation statements)

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“…The temperature dependence of the excess heat capacity was further analyzed and plotted using Origin 7.0 software (MicroCal Inc., Northampton, MA, USA). The deconvolution procedure was carried out in the Origin 7.0 software using the algorithms described earlier [48,49].…”

Section: Differential Scanning Calorimetry (Dsc)mentioning

confidence: 99%

Impact of A134 and E218 Amino Acid Residues of Tropomyosin on Its Flexibility and Function

Marchenko

Nefedova

Yampolskaya

et al. 2020

IJMS

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Tropomyosin (Tpm) is one of the major actin-binding proteins that play a crucial role in the regulation of muscle contraction. The flexibility of the Tpm molecule is believed to be vital for its functioning, although its role and significance are under discussion. We choose two sites of the Tpm molecule that presumably have high flexibility and stabilized them with the A134L or E218L substitutions. Applying differential scanning calorimetry (DSC), molecular dynamics (MD), co-sedimentation, trypsin digestion, and in vitro motility assay, we characterized the properties of Tpm molecules with these substitutions. The A134L mutation prevented proteolysis of Tpm molecule by trypsin, and both substitutions increased the thermal stability of Tpm and its bending stiffness estimated from MD simulation. None of these mutations affected the primary binding of Tpm to F-actin; still, both of them increased the thermal stability of the actin-Tpm complex and maximal sliding velocity of regulated thin filaments in vitro at a saturating Ca2+ concentration. However, the mutations differently affected the Ca2+ sensitivity of the sliding velocity and pulling force produced by myosin heads. The data suggest that both regions of instability are essential for correct regulation and fine-tuning of Ca2+-dependent interaction of myosin heads with F-actin.

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Section: Differential Scanning Calorimetry (Dsc)mentioning

confidence: 99%

Impact of A134 and E218 Amino Acid Residues of Tropomyosin on Its Flexibility and Function

Marchenko

Nefedova

Yampolskaya

et al. 2020

IJMS

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“…22 These our data are similar to those obtained earlier for αα-Tpm homodimers carrying cardiomyopathy-associated mutation M8R which also did not affect the thermal stability of Tpm, but sharply decreased the viscosity of Tpm solution and the affinity of αα-Tpm for F-actin. 41 The N-terminal sequences of more than 20 residues containing Met9 (or Met8 in α-Tpm) residue are almost identical for γ-Tpm and α-Tpm, except only substitution of Asp2 in α-Tpm with Glu2 in γ-Tpm and the presence of additional Met residue at the N-terminus of γ-Tpm. 6 The conserved Met9 residues in WT Tpm participate in the formation of hydrophobic interactions between the N-and C-termini of neighboring Tpm molecules.…”

Section: Effects Of the M9r Mutationmentioning

confidence: 92%

“…Previous studies indicated that the stability of the Tpm-F-actin complex depends, to a great extent, on the thermal stability of the Tpm molecule rather than on the affinity of Tpm for F-actin. 35,37,39,41 This explains why the K169E substitution, which strongly destabilized the γγ-Tpm molecule (Figure 2), also destabilized its complex with F-actin ( Figure 5A). In all Tpm dimers (γ*γ*-, γ*β-, and γ*γ-Tpm), the K169E substitution decreased the sliding velocity of the filaments irrespective of the presence or absence of Tn and Ca 2+ ( Figure 6, Abbreviations: Fr 0 (%), fraction of motile filaments at low Ca 2+ concentration (pCa 7.5-8.0); Fr max (%), fraction of motile filaments at high Ca 2+ concentration (pCa 5); pCa 50 , pCa at which the fraction of filaments moving is halfmaximal;γ*β-Tpm, γβ-heterodimer of Tpm with mutation in the γ-chain; γ*γ*-Tpm, γγ-homodimer of Tpm with mutations in both γ-chains; γ*γ-Tpm, γγ-homodimer of Tpm with mutation in only one γ-chain.…”

Section: Effects Of the K169e Mutationmentioning

confidence: 95%

Mechanisms of disturbance of the contractile function of slow skeletal muscles induced by myopathic mutations in the tropomyosinTPM3gene

et al. 2020

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Several congenital myopathies of slow skeletal muscles are associated with mutations in the tropomyosin (Tpm) TPM3 gene. Tropomyosin is an actin‐binding protein that plays a crucial role in the regulation of muscle contraction. Two Tpm isoforms, γ (Tpm3.12) and β (Tpm2.2) are expressed in human slow skeletal muscles forming γγ‐homodimers and γβ‐heterodimers of Tpm molecules. We applied various methods to investigate how myopathy‐causing mutations M9R, E151A, and K169E in the Tpm γ‐chain modify the structure‐functional properties of Tpm dimers, and how this affects the muscle functioning. The results show that the features of γγ‐Tpm and γβ‐Tpm with substitutions in the Tpm γ‐chain vary significantly. The characteristics of the γγ‐Tpm depend on whether these mutations located in only one or both γ‐chains. The mechanism of the development of nemaline myopathy associated with the M9R mutation was revealed. At the molecular level, a cause‐and‐effect relationship has been established for the development of myopathy by the K169E mutation. Also, we described the structure‐functional properties of the Tpm dimers with the E151A mutation, which explain muscle weakness linked to this substitution. The results demonstrate a diversity of the molecular mechanisms of myopathy pathogenesis induced by studied Tpm mutations.

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“…All Tpm constructs were expressed in E. coli C41(DE3) cells by standard protocol, and then extracted and purified by anion exchange chromatography using a HiTrap QHP column (GE Healthcare, Marlborough, MA, USA) as previously described [ 30 , 41 ]. Tpm concentration was determined spectrophotometrically at 280 nm using an E 1% of 2.7 cm −1 for Tpm1.5, 1.8 cm −1 for Tpm1.6 and Tpm1.7, 2.1 cm −1 for Tpm1.12, and 1.6 cm −1 for Tpm4.2.…”

Section: Methodsmentioning

confidence: 99%

“…The thermal stability of the proteins was described by the temperature of the maximum of the thermal transition (T m ), and the calorimetric enthalpy (ΔH cal ) was calculated as the area under the excess heat capacity function. Deconvolution analysis of the heat sorption curves, that is, their decomposition into separate thermal transitions (calorimetric domains) by fitting the data to the non-two-state model [ 43 ], was performed as described earlier [ 28 , 31 , 33 , 41 , 44 ].…”

Section: Methodsmentioning

confidence: 99%

Structural and Functional Peculiarities of Cytoplasmic Tropomyosin Isoforms, the Products of TPM1 and TPM4 Genes

Marchenko

Nefedova

Artemova

et al. 2021

IJMS

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Tropomyosin (Tpm) is one of the major protein partners of actin. Tpm molecules are α-helical coiled-coil protein dimers forming a continuous head-to-tail polymer along the actin filament. Human cells produce a large number of Tpm isoforms that are thought to play a significant role in determining actin cytoskeletal functions. Even though the role of these Tpm isoforms in different non-muscle cells is more or less studied in many laboratories, little is known about their structural and functional properties. In the present work, we have applied various methods to investigate the properties of five cytoplasmic Tpm isoforms (Tpm1.5, Tpm 1.6, Tpm1.7, Tpm1.12, and Tpm 4.2), which are the products of two different genes, TPM1 and TPM4, and also significantly differ by alternatively spliced exons: N-terminal exons 1a2b or 1b, internal exons 6a or 6b, and C-terminal exons 9a, 9c or 9d. Our results demonstrate that structural and functional properties of these Tpm isoforms are quite different depending on sequence variations in alternatively spliced regions of their molecules. The revealed differences can be important in further studies to explain why various Tpm isoforms interact uniquely with actin filaments, thus playing an important role in the organization and dynamics of the cytoskeleton.

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The effects of cardiomyopathy-associated mutations in the head-to-tail overlap junction of α-tropomyosin on its properties and interaction with actin

Cited by 21 publications

References 50 publications

Impact of A134 and E218 Amino Acid Residues of Tropomyosin on Its Flexibility and Function

Impact of A134 and E218 Amino Acid Residues of Tropomyosin on Its Flexibility and Function

Mechanisms of disturbance of the contractile function of slow skeletal muscles induced by myopathic mutations in the tropomyosinTPM3gene

Structural and Functional Peculiarities of Cytoplasmic Tropomyosin Isoforms, the Products of TPM1 and TPM4 Genes

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