William C.T. Yang

Formation of creatine phosphate from creatine and 32P-labelled ATP by isolated rabbit heart mitochondria.

Abstract With ATP [γ- 32 P] we have demonstrated directly that mitochondrial creatine phosphokinase catalyzes the formation of large amounts of creatine phosphate with mitochondria generated ATP as substrate rather than added extramitochondrial ATP.

Intimate coupling of creatine phosphokinase and myofibrillar adenosinetriphosphatase

Abstract ATPase and creatine phosphokinase (CPK) activities of isolated cardiac myofibrils were determined with 32 P γ-labeled ATP alone and with the addition of phosphorylcreatine (PC). With ATP and PC as substrates the label in the inorganic phosphate formed is greatly diluted indicating that the ATP formed by PC through CPK can reach the ATPase active site more readily than labeled ATP from the medium. The tight coupling of the ATPase and CPK activities further strengthens our view that PC serves an important role as high energy carrier between the energy producing sites (mitochondria) and the energy utilizing sites (myofibrils).

Mechanism of the inhibitory action of isoniazid on microsomal drug metabolism

Abstract Addition of isoniazid (isonicotinic acid hydrazide, INH) to rat liver microsomes produced an immediate decrease in the binding of carbon monoxide to reduced cytochrome P-450. Preincubation of the microsomes with INH in the presence of NADPH produced a further decrease of carbon monoxide binding to cytochrome P-450. The latter decrease of functional cytochrome P-450 was dependent upon NADPH and oxygen and was transitory. Examination of compounds structurally related to INH indicated that both the hydrazine moiety and the aromatic ring were needed to produce both effects. Incubation of microsomes with INH also resulted in gradual increases in absorbance at 449 nm and at 493 nm which also were transitory. Thus, the decreased binding of carbon monoxide to cytochrome P-450 may have occurred concurrently with formation of these spectral intermediates. Microsomal N -demethylation and aniline p -hydroxylation were inhibited by isoniazid. Preincubation of the microsomes with INH and NADPH increased the inhibition. Thus, the decreased availability of cytochrome P-450 as observed may account for the inhibition of the mixed function oxidases by isoniazid.

Inhibition of cardiac creatine phosphokinase by fluorodinitrobenzene

Abstract With electrophoretic evidence for a mitochondrial isoenzyme of creatine phosphokinase (CPK), we feel that functional studies are necessary to help further elucidate the properties of this isoenzyme. As one approach, fluorodinitrobenzene (FDNB) was used to examine its effect on mitochondrial CPK. In both polarographic studies and direct enzymatic studies, 10 −5 M FDNB was shown to almost completely inhibit the enzyme activity, as has been shown in skeletal muscle. In addition it was observed that the mitochondrial CPK was just as susceptible to the inhibitory effect of FDNB as the cytoplasmic isoenzyme.

The Creatine-Creatine Phosphate Shuttle for Energy Transport — Compartmentation of Creatine Phosphokinase in Muscle

There are two viewpoints regarding the role of the enzyme creatine Phosphokinase (CPK), which catalyzes the reversible transphosphorylation of creatine and adenine nucleotides in energy transport. The classical viewpoint (Fig. 1) regards CPK as a near equilibrium enzyme buffering cytosolic changes in nucleotide concentration (1,2), with creatine phosphate representing a reservoir of high energy phosphate equivalents (3) and with regulation of energy production relegated to the adenine nucleotides whether in the form of energy charge (4), adenylate phosphate potential (5,6), or ATP/ADP ratio (7). The second viewpoint places CPK and its products, creatine and creatine phosphate, in the central role of energy transport (8–11). The evidence for the latter viewpoint has come about from several diverse lines of reasoning (reviewed in 9). Jacobs et al discovered a distinct isoenzyme of CPK bound to mitochondria in 1964 (12). The significance of this mitochondrial bound form of CPK was studied by Bessman and Fonyo (13) in pigeon breast mitochondria where the possibility of feedback regulation of respiration through production of acceptor creatine was suggested, in accordance with Bessman’s earlier observation on the insulin-like effect of exercise in diabetics (14).

COMPARATIVE ACTIONS OF 1,10-PHENANTHROLINE AND QUINIDINE ON NORMAL AND FIBRILLATING ISOLATED ATRIA.

Summary 1. A method of inducing fibrillation in isolated rabbit atria was explored. Atrial fibrillation can be readily produced and maintained under the conditions described by Holland and Burn(2). The method is improved here by the use of much lower concentration of acetylcholine. 2. 1,10-Phenanthroline is as effective as quinidine in stopping induced fibrillation in isolated rabbit atria. 3. 1,10-Phenanthroline, like quinidine, reduces the maximal following rate of isolated rabbit atria, but to a lesser extent. 4. The lowering of maximal following rate by 1,10-Phenanthroline and quinidine can be lessened by reduction of potassium in the medium. 5. At comparable concentrations, 1,10-Phenanthroline does not affect the intrinsic sinus rate of isolated rabbit atria, while quinidine reduces the intrinsic rate markedly. 6. 1,10-Phenanthroline, like quinidine, produces distinct reduction of the rate of depolarization and repolarization of the action potential of the rat atria.