J. Biol. Chem., Vol. 261, Issue 26, 12141-12146, 09, 1986
The two pathways for oxygen exchange by actomyosin and myofibrils and their dependence on temperature
KK Shukla, HM Levy, F Ramirez and JF Marecek
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.
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