Vincent S H, Silverman D N
J Biol Chem. 1982 Jun 25;257(12):6850-5.
An 18O exchange method has been used to determine the location of carbonic anhydrase in mitochondria from rat liver and to examine the role of this enzyme in the kinetics of CO2 in resting and respiring mitochondria. Using digitonin subfractionation, we have determined that a substantial fraction, 40 to 60%, of the carbonic anhydrase activity in the mitochondrion from rat liver is located in the space between the inner and outer membranes; the remaining activity was found in the matrix with no detectable activity in the sedimented membranes. The total catalytic CO2 hydration activity measured in intact mitochondria from rat liver was about 1% of that found in an equal volume of rat erythrocytes. The apparent permeability constant representing the barrier for the diffusion of HCO3(-) from external solution to intramitochondrial carbonic anhydrase, 9 X 10(-5) cm s-1, is near in magnitude to the permeability constant for the diffusion of HCO3(-) across the rat erythrocyte membrane, 4 X 10(-4) cm s-2. Calcium-induced respiratory jumps were shown to cause changes in the rate of 18O exchange between CO2 and H2O that were consistent with a net uptake of CO2 by the mitochondria.
采用18O交换法来确定大鼠肝脏线粒体中碳酸酐酶的位置，并研究该酶在静息和呼吸状态下的线粒体中二氧化碳动力学方面的作用。利用洋地黄皂苷分级分离法，我们已确定大鼠肝脏线粒体中40%至60%的碳酸酐酶活性位于内膜和外膜之间的空间；其余活性存在于基质中，沉淀的膜中未检测到活性。在大鼠肝脏完整线粒体中测得的总催化二氧化碳水合活性约为等量大鼠红细胞中活性的1%。代表HCO3(-)从外部溶液扩散到线粒体内碳酸酐酶的屏障的表观渗透常数为9×10(-5) cm s-1，其大小与HCO3(-)跨大鼠红细胞膜扩散的渗透常数4×10(-4) cm s-2相近。钙诱导的呼吸跃变被证明会导致CO2与H2O之间18O交换速率的变化，这与线粒体对CO2的净摄取一致。
