The Journal of General Physiology
Cell MicroControls
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The Journal of General Physiology, Vol 48, 737-752, Copyright © 1965 by The Rockefeller University Press

ARTICLE

Isolation of Relaxing Particles from Rat Skeletal Muscles in Zonal Centrifuges

H. Schuel 1, L. Lorand 1, R. Schuel 1, and N. G. Anderson 1

1 From the Department of Chemistry, Northwestern University, Evanston, Illinois, and Biology Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee

Supernatants of rat skeletal muscle homogenates were fractionated by differential centrifugation and by zonal centrifugation in sucrose density gradients. Cytochrome oxidase was employed as an enzymatic marker for locating mitochondria. The subcellular fractions were also assayed for their ability to prevent the ATP-induced contraction of myofibrils. Both the mitochondrial and microsomal fractions obtained by differential fractionation were found to be rich in such relaxing activity, and the microsomal fraction was appreciably contaminated by mitochondria. In contrast to this, when fractionation was carried out by means of zonal centrifugation (4200 RPM x 205 min. to 40,000 RPM x 60 min.), relaxing activity was found to be associated only with particles having the sedimentation characteristics of microsomes (s20,w estimated to be between 370 and 1880S). Relaxing activity was not detected in the regions of the gradient containing either the starting sample zone (soluble phase) or the mitochondrial peak. The microsomal relaxing particles showed negligible cytochrome oxidase activity.

Submitted on November 9, 1964


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N. G. Anderson
Zonal Centrifuges and Other Separation Systems
Science, October 7, 1966; 154(3745): 103 - 112.
[Abstract] [PDF]


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