Characteristics of Sarcoplasmic Reticulum Membrane Preparations Isolated from Skeletal Muscles of Active and Hibernating Ground Squirrel Spermophilus undulatus

A. N. Malysheva¹, K. B. Storey², R. Kh. Ziganshin³, O. D. Lopina¹, and A. M. Rubtsov^1*

¹Department of Biochemistry, School of Biology, Lomonosov Moscow State University, Moscow, 119899 Russia; fax: (095) 939-3955; E-mail: am_rubtsov@mail.ru

²Department of Biology and Department of Chemistry, Carleton University, Ottawa K1S 5B6, Canada; fax: (613) 520-2569; E-mail: kbstorey@ccs.carleton.ca

³Laboratory of Peptide Chemistry, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, 117871 Russia; fax: (095) 335-7103; E-mail: rustam@ibch.siobc.rac.ru

^* To whom correspondence should be addressed.

Received January 25, 2001; Revision received February 20, 2001

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The total Ca-ATPase activity in the sarcoplasmic reticulum (SR) membrane fraction isolated from skeletal muscles of winter hibernating ground squirrel Spermophilus undulatus is ~2.2-fold lower than in preparations obtained from summer active animals. This is connected in part with ~10% decrease of the content of Ca-ATPase protein in SR membranes. However, the enzyme specific activity calculated with correction for its content in SR preparations is still ~2-fold lower in hibernating animals. Analysis of the protein composition of SR membranes has shown that in addition to the decrease in Ca-ATPase content in hibernating animals, the amount of SR Ca-release channel (ryanodine receptor) is decreased ~2-fold, content of Ca-binding proteins calsequestrin, sarcalumenin, and histidine-rich Ca-binding protein is decreased ~3-4-fold, and the amount of proteins with molecular masses 55, 30, and 22 kD is significantly increased. Using the cross-linking agent cupric-phenanthroline, it was shown that in SR membranes of hibernating ground squirrels Ca-ATPase is present in a more aggregated state. The affinity of SR membranes to the hydrophilic fluorescent probe ANS is higher and the degree of excimerization of the hydrophobic probe pyrene is lower (especially for annular lipids) in preparations from hibernating than from summer active animals. The latter indicates an increase in the microviscosity of the lipid environment of Ca-ATPase during hibernation. We suggest that protein aggregation as well as the changes in protein composition and/or in properties of lipid bilayer SR membranes can result in the decrease of enzyme activity during hibernation.

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KEY WORDS: sarcoplasmic reticulum, Ca-ATPase, Ca-binding proteins, calsequestrin, sarcalumenin, histidine-rich Ca-binding protein, ground squirrel Spermophilus undulatus

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