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The two pathways for oxygen exchange by actomyosin and myofibrils and their dependence on temperature
Abstract:
At an intermediate stage in the hydrolysis of MgATP by actomyosin there is an exchange of oxygen between water and the terminal phosphoryl group of MgATP, tightly bound to the myosin active site. This intermediate oxygen exchange results from the reversible hydrolysis of the bound MgATP. The rate of the exchange cycle (hydrolysis and the reverse) is assumed to be determined by the rate of reverse hydrolysis; and the average time available for exchange is determined by the post-exchange reaction that immediately follows the cycle. Past analytical studies of the exchange, using actomyosin mixtures and myofibrils at room temperature, have revealed two pathways for hydrolysis, operating at a comparable flux but differing greatly in the extent of exchange they support. It is shown here that these pathways also appear over a range of temperatures from 5 to 30 degrees C and that temperature had little effect on their relative fluxes. At each temperature, the flux ratio (%) for the low exchange pathway: high exchange pathway was near 50:50 for actomyosin mixtures and 60:40 for myofibrils. Apparently, the rate-limiting steps that determine the fluxes of the two pathways have a similar temperature dependence. However, the analysis indicates that one or both of the steps that determine the extent of exchange (reverse-hydrolysis and/or the post-exchange reaction) shows a different temperature dependence for the two pathways. We interpret this to reflect a difference in the temperature dependence of the post-exchange reaction, which we propose is exceedingly fast and independent of actin concentration along the low exchange route, but slow and dependent on the actin concentration along the high exchange route. Thus at all temperatures over a broad range of actin concentration there are two pathways of comparable flux that differ primarily in the time available for exchange.
Insights
Two MgATP hydrolysis pathways in actomyosin show similar temperature-dependent fluxes but differ in oxygen exchange extent. This difference is attributed to the post-exchange reaction
Area of Science:
- Biochemistry
- Muscle Physiology
Background:
- MgATP hydrolysis by actomyosin involves oxygen exchange between water and MgATP.
- This exchange occurs at an intermediate stage and results from reversible MgATP hydrolysis.
- Two distinct pathways for MgATP hydrolysis have been previously identified, differing in exchange extent.
Purpose of the Study:
- To investigate the temperature dependence of MgATP hydrolysis pathways.
- To understand how temperature affects the relative fluxes and oxygen exchange extents of these pathways.
- To elucidate the mechanisms underlying the differences in exchange extent between the pathways.
Main Methods:
- Analysis of oxygen exchange during MgATP hydrolysis by actomyosin and myofibrils.
- Experiments conducted over a temperature range of 5 to 30 degrees C.
- Comparison of flux ratios and exchange extents between low and high exchange pathways.
Main Results:
- Both low and high oxygen exchange pathways persist across the tested temperature range (5-30°C).
- Temperature had minimal impact on the relative fluxes of the two pathways.
- Flux ratios were approximately 50:50 for actomyosin and 60:40 for myofibrils.
- The extent of exchange differs significantly between pathways, suggesting varying temperature dependencies in rate-limiting steps.
Conclusions:
- MgATP hydrolysis occurs via two pathways with comparable fluxes but differing exchange extents.
- The post-exchange reaction's temperature dependence likely explains the differing exchange extents between pathways.
- The low exchange pathway features a fast, actin-independent post-exchange reaction, while the high exchange pathway has a slow, actin-dependent one.