Differential scanning calorimetry study of glycerinated rabbit psoas muscle fibres in intermediate state of ATP hydrolysisReportar como inadecuado

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BMC Structural Biology

, 7:41

First Online: 24 June 2007Received: 03 August 2006Accepted: 24 June 2007


BackgroundThermal denaturation experiments were extended to study the thermal behaviour of the main motor proteins actin and myosin in their native environment in striated muscle fibres. The interaction of actin with myosin in the highly organized muscle structure is affected by internal forces; therefore their altered conformation and interaction may differ from those obtained in solution. The energetics of long functioning intermediate states of ATP hydrolysis cycle was studied in muscle fibres by differential scanning calorimetry DSC.

ResultsSETARAM Micro DSC-II was used to monitor the thermal denaturation of the fibre system in rigor and in the presence of nucleotide and nucleotide analogues. The AM.ADP.Pi state of the ATP hydrolysis cycle has a very short lifetime therefore, we mimicked the different intermediate states with AMP.PNP and-or inorganic phosphate analogues Vi and AlF4 or BeFx. Studying glycerol-extracted muscle fibres from the rabbit psoas muscle by DSC, three characteristic thermal transitions were detected in rigor. The thermal transitions can be assigned to myosin heads, myosin rods and actin with transition temperatures Tm of 52.9 ± 0.7°C, 57.9 ± 0.7°C, 63.7 ± 1.0°C. In different intermediate states of the ATP hydrolysis mimicked by nucleotide analogues a fourth thermal transition was also detected which is very likely connected with nucleotide binding domain of myosin and-or actin filaments. This transition temperature Tm4 depended on the mimicked intermediate states, and varied in the range of 66°C – 77°C.

ConclusionAccording to DSC measurements, strongly and weakly binding states of myosin to actin were significantly different. In the presence of ADP only a moderate change of the DSC pattern was detected in comparison with rigor, whereas in ADP.Pi state trapped by Vi, AlF4 or BeFx a remarkable stabilization was detected on the myosin head and actin filament which is reflected in a 3.0 – 10.0°C shift in Tm to higher temperature. A similar effect was observed in the case of the nonhydrolyzable AMP.PNP analogue. Differential DSC measurements suggest that stabilization actin structure in the intermediate states of ATP hydrolysis may play an additional role in actin-myosin interaction.

Electronic supplementary materialThe online version of this article doi:10.1186-1472-6807-7-41 contains supplementary material, which is available to authorized users.

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Autor: Timea Dergez - Dénes Lőrinczy - Franciska Könczöl - Nelli Farkas - Joseph Belagyi

Fuente: https://link.springer.com/

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