Oxidative phosphorylation in insect sarcosomes

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Abstract
Oxidative phosphorylation was demonstrated with sarcosomes isolated from the thoracic muscle of the blowfly Calliphora erythrocephala. The P:O ratio varied considerably from preparation to preparation, but never exceeded 2.0. The ratio was lower with sarcosomes isolated from young flies. Factors causing low ratios were (1) isolation in media hypotonic or hypertonic to the blowfly hemolymph (tonicity equal to that of 0.36M sucrose); (2) isolation in potassium chloride or phosphate; (3) allowing the dissected thoraces to stand at room temperature for 35 minutes; (4) subjecting the thoraces to more than gentle crushing to liberate the sarcosomes; (5) omission of ethylenediaminetetraacetic acid from the reaction mixture; (6) prolonging the experiment beyond 35 minutes (at 25[degree]). The treatments referred to in (1) and (2) gave preparations with higher oxidative activity. The inclusion of fluoride or malonate in the reaction mixture or variation of the alpha-ketoglutarate concentration between 0.002 and 0.01M had no effect on the phosphorylation ratios. Sarcosomes isolated in sucrose containing ethylenediaminetetraacetic acid oxidized alpha-ketoglutarate more slowly than those isolated in sucrose alone. The P:O ratio was little affected. The addition of small amounts of albumin (1.2 mg/ml) to the reaction mixture markedly stimulated the rate of oxidation of alpha-ketoglutarate, without usually affecting the phosphorylation ratio. Albumin considerably increased the P:O ratio when succinate was the substrate. Albumin is a true activator of the oxidase system, in contrast with ethylenediaminetetraacetic acid which stabilizes the system. Apart from the activation caused by the introduction of albumin into the reaction mixture, the inclusion of albumin in the isolation medium had little effect.