EXAMINATION OF PLASMA MEMBRANE-ENRICHED FRACTION FROM GUINEA PIG INTESTINAL SMOOTH MUSCLE BY MEANS OF SOME MARKER ENZYMES

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By means of a preparation technique based on the discontinuous sucrose density gradient, subcellular fractions were isolated from guinea pig intestinal smooth muscle cells. A fraction which distributed to a 33% sucrose layer showed relatively high activities of 5'-nucleotidase, Na+·K+-ATPase and ouabain sensitive Na+·K+-ATPase. The fraction had a low NaN3 sensitive Mg2+-ATPase activity. On the other hand, the high activity of glucose-6-phosphatase showed a broad distribution. Though the sucrose density gradient proceeded over a series of fine layers, cross-contamination of microsome into the 33% sucrose fraction was not reduced. To reduce microsomal cross-contamination, another procedure was employed. The homogenization time of 77000×g sediment to be layered on the top of the sucrose density gradients was prolonged. This procedure did not change the distribution of K+ activated p-nitrophenylphosphatase, K+ activated ouabain sensitive p-nitrophenylphosphatase and ouabain sensitive Na+. K+-ATPase activities. The peak of NADH cytochrome c reductase activity was shifted to a 38% sucrose fraction from a 33% sucrose fraction and the activity of this marker enzyme in the 33% sucrose fraction decreased to 60% of that of the prior procedure.
日本平滑筋学会の論文