title: Inhibition of sarcoplasmic reticulum calcium pump by cytosolic protein(s) endogenous to heart and slow skeletal muscle but not fast skeletal muscle.
abstract: Cytosol from rabbit heart and slow and fast skeletal muscles was fractionated using (NH4)2SO4 to yield three cytosolic protein fractions, viz., CPF-I (protein precipitated at 30% saturation), CPF-II (protein precipitated between 30 and 60% saturation), and cytosol supernatant (protein soluble at 60% saturation). The protein fractions were dialysed and tested for their effects on ATP-dependent, oxalate-supported Ca2+ uptake by sarcoplasmic reticulum from heart and slow and fast skeletal muscles. CPF-I from heart and slow muscle, but not from fast muscle, caused marked inhibition (up to 95%) of Ca2+ uptake by sarcoplasmic reticulum from heart and from slow and fast muscles. Neither unfractionated cytosol nor CPF-II or cytosol supernatant from any of the muscles altered the Ca2+ uptake activity of sarcoplasmic reticulum. Studies on the characteristics of inhibition of sarcoplasmic reticulum Ca2+ uptake by CPF-I (from heart and slow muscle) revealed the following: (a) Inhibition was concentration- and temperature-dependent (50% inhibition with approx. 80 to 100 micrograms CPF-I; seen only at temperatures above 20 degrees C). (b) The inhibitor reduced the velocity of Ca2+ uptake without appreciably influencing the apparent affinity of the transport system for Ca2+. (c) Inhibition was uncompetitive with respect to ATP. (d) Sarcoplasmic reticulum washed following exposure to CPF-I showed reduced rates of Ca2+ uptake, indicating that inhibition results from an interaction of the inhibitor with the sarcoplasmic reticulum membrane. (e) Concomitant with the inhibition of Ca2+ uptake, CPF-I also inhibited the Ca2+-ATPase activity of sarcoplasmic reticulum. (f) Heat-treatment of CPF-I led to loss of inhibitor activity, whereas exposure to trypsin appeared to enhance its inhibitory effect. (g) Addition of CPF-I to Ca2+-preloaded sarcoplasmic reticulum vesicles did not promote Ca2+ release from the vesicles. These results demonstrate the presence of a soluble protein inhibitor of sarcoplasmic reticulum Ca2+ pump in heart and slow skeletal muscle but not in fast skeletal muscle. The characteristics of the inhibitor and its apparently selective distribution suggest a potentially important role for it in the in vivo regulation of sarcoplasmic reticulum Ca2+ pump, and therefore in determining the duration of Ca2+ signal in slow-contracting muscle fibers.
mesh_terms: D000818:Animals; D001693:Biological Transport, Active; D002122:Calcium Chloride / Q000378:metabolism; D003600:Cytosol / Q000502:physiology; D006321:Heart / Q000502:physiology*; D007473:Ion Channels / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D009124:Muscle Proteins / Q000302:isolation & purification / Q000502:physiology*; D009132:Muscles / Q000502:physiology*; D009928:Organ Specificity; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
chemical_list: D007473:Ion Channels; D009124:Muscle Proteins; D002122:Calcium Chloride