The Optimized Quantum Dots Mediated Thermometry Reveals the Efficiency of Myosin Extracted from Muscle Mini Bundles
Abstract Background: The quantum dots (QD) has been investigated as thermometrical sensor in biological microenvironment and applied to measure the muscle efficiency with underlying mechanisms, i.e., a reduction in fluorescent intensity of QD reflects an increase in temperature caused by heat release during ATP hydrolysis, denoting the efficiency of the motor protein myosin. The aim of this study is to optimize the QD mediated thermometry for measuring the efficiency of freshly extracted myosin from muscle mini bundles rather than pre-purified myosin and test this approach in preparations with different myosin isoform.Methods: The protocol of myosin extraction used in the single muscle fiber in vitro motility assay was modified slightly for extracting myosin from the muscle mini bundles. Moreover, the quantitation of extracted myosin was calculated from the total extracted protein since the ratio of myosin to total protein was constant, performing through spectrophotometric measurement of UV absorbance at 280 nm. The change in fluorescence intensity of QD thermometry measurement of myosin ATPase enzymatic reaction was plotted over time, and the slope of the linear negative regression between time course and relatively decreased fluorescence intensity was used to reflect the efficiency of extracted myosin. Results: The optimized QD mediated thermometry is established for evaluating the efficiency of myosin extracted from muscle mini bundles. Moreover, myosin isoform specific differences in the myosin efficiency were observed in comparison between slow myosin and fast myosin, i.e., the low myosin has lower efficiency than fast myosin, evidenced by a higher heat release.Conclusions: The optimized QD mediated thermometric measure of myosin efficiency in muscle mini bundles provides a nanoscale approach to evaluate myosin function based on a minimal amount of muscle, which is essentially required in the muscle research..
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Preprint| Erscheinungsjahr: |
2021 |
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| Erschienen: |
2021 |
| Enthalten in: |
ResearchSquare.com - (2021) vom: 22. Sept. Zur Gesamtaufnahme - year:2021 |
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| Sprache: |
Englisch |
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| Beteiligte Personen: |
Li, Meishan [VerfasserIn] |
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| Links: |
Volltext [kostenfrei] |
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| doi: |
10.21203/rs.3.rs-903082/v1 |
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XRA034755993 |
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| 520 | |a Abstract Background: The quantum dots (QD) has been investigated as thermometrical sensor in biological microenvironment and applied to measure the muscle efficiency with underlying mechanisms, i.e., a reduction in fluorescent intensity of QD reflects an increase in temperature caused by heat release during ATP hydrolysis, denoting the efficiency of the motor protein myosin. The aim of this study is to optimize the QD mediated thermometry for measuring the efficiency of freshly extracted myosin from muscle mini bundles rather than pre-purified myosin and test this approach in preparations with different myosin isoform.Methods: The protocol of myosin extraction used in the single muscle fiber in vitro motility assay was modified slightly for extracting myosin from the muscle mini bundles. Moreover, the quantitation of extracted myosin was calculated from the total extracted protein since the ratio of myosin to total protein was constant, performing through spectrophotometric measurement of UV absorbance at 280 nm. The change in fluorescence intensity of QD thermometry measurement of myosin ATPase enzymatic reaction was plotted over time, and the slope of the linear negative regression between time course and relatively decreased fluorescence intensity was used to reflect the efficiency of extracted myosin. Results: The optimized QD mediated thermometry is established for evaluating the efficiency of myosin extracted from muscle mini bundles. Moreover, myosin isoform specific differences in the myosin efficiency were observed in comparison between slow myosin and fast myosin, i.e., the low myosin has lower efficiency than fast myosin, evidenced by a higher heat release.Conclusions: The optimized QD mediated thermometric measure of myosin efficiency in muscle mini bundles provides a nanoscale approach to evaluate myosin function based on a minimal amount of muscle, which is essentially required in the muscle research. | ||
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