Does pressure antagonize anesthesia? Opposite effects on specific and nonspecific inhibitors of firefly luciferase

Ueda and Suzuki (1998. Biochim. Biophys. Acta. 1380:313-319; 1998. Biophys. J. 75:1052-1057) reported that myristic acid inhibited firefly luciferase in mu M range in competition with luciferin, whereas anesthetics inhibited it in millimeter ranges noncompetitively with luciferin. Myristate increase d, whereas anesthetics decreased, the thermal denaturation temperature. The present study showed that high pressure increased the steady-state light i ntensity of the halothane-doped firefly luciferase but decreased that of th e myristate-doped firefly luciferase. The steady-state light intensity show ed a maximum at 19.1 degrees C. At 19.1 degrees C, high pressure did not af fect the light intensity in the absence of the inhibitors. In the presence of 0.5 mM halothane, however, 25 MPa pressure (maximum effect) increased th e light intensity to 106.0% of the control without the inhibitor. In the pr esence of 2.5 mu M myristate, 40 MPa pressure decreased the light intensity to 90.9% of the control. When the temperature was 25 degrees C in the abse nce of inhibitors, 40 MPa pressure increased the tight intensity 119.2% of the ambient value. At 0.5 mM halothane, 40 MPa pressure further increased t he light intensity to 106.1% above the control 40 MPa value. At 2.5 mu M my ristate, 40 MPa pressure decreased the light intensity to 90.1% of the cont rol 40 MPa value. From the pressure dependence of the light intensity, the volume change Delta V of the enzyme was estimated at 25 degrees C: 0.5 mM h alothane increased Delta V = +3.93 cm(3) mol(-1), whereas 2.5 mu M myristat e decreased Delta V = -7.66 cm(3) mol(-1). Present results show that there are distinct differences between the specific and nonspecific ligands in th eir response to high pressure. Myristate, which competes with luciferin, de creased the protein volume and stabilized the conformation against thermal perturbation. Halothane, which does not compete with the substrate, increas ed the protein volume and destabilized the conformation.