QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online January 31, 2006
Journal of Experimental Biology 209, 677-688 (2006)
Published by The Company of Biologists 2006
doi: 10.1242/jeb.02052

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Ramnanan, C. J. Articles by Storey, K. B. Search for Related Content PubMed PubMed Citation Articles by Ramnanan, C. J. Articles by Storey, K. B.

Suppression of Na⁺/K⁺-ATPase activity during estivation in the land snail Otala lactea

Christopher J. Ramnanan and Kenneth B. Storey^*

Institute of Biochemistry and Department of Biology, College of Natural Sciences, Carleton University, 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S 5B6

View larger version (16K): [in a new window]   Fig. 1. Na+/K+-ATPase activity in hepatopancreas from active and estivated O. lactea as a function of assay pH. Assays were conducted under optimal substrate concentrations at 22°C. Data are means ± s.e.m., N=4 independent preparations of enzyme.

View larger version (17K): [in a new window]   Fig. 2. Na+/K+-ATPase activity in O. lactea (A) foot muscle (mean ± s.e.m., N=9) and (B) hepatopancreas (mean ± s.e.m., N=4) as a function of Mg.ATP concentration.

View larger version (19K): [in a new window]   Fig. 3. Substrate affinity for ATP as a function of temperature for Na+/K+-ATPase activity in O. lactea in (A) foot muscle and (B) hepatopancreas. Values are means ± s.e.m., N=4 independent preparations of enzyme except for foot muscle control and estivated activity, which are N=9. *Significantly different from the corresponding control incubation; Student's t-test, P<0.01.

View larger version (30K): [in a new window]   Fig. 4. Effect of in vitro incubations to stimulate the activities of endogenous (A) protein kinases and (B) protein phosphatases on Na+/K+-ATPase activity in foot muscle extracts from active and 10-day estivated O. lactea. Incubation time was 4 h prior to assay under Vmax conditions at 22°C. Values are means ± s.e.m., N=4 independent determinations. *Significantly different from the corresponding control incubation; Student's t-test, P<0.01; P<0.05. For details of incubation conditions, see Materials and methods.

View larger version (30K): [in a new window]   Fig. 5. Effect of in vitro incubations stimulating the activities of endogenous (A) protein kinases and (B) protein phosphatases on Na+/K+-ATPase activity in hepatopancreas extracts from active and 10-day estivated O. lactea. Other information as in Fig. 4.

View larger version (21K): [in a new window]   Fig. 6. Effect of calcium concentration on the stimulation by endogenous protein kinases of Na+/K+-ATPase activity in foot muscle extracts from active snails. In vitro incubations stimulated PKA, PKC or PKG and the resulting change in Na+/K+-ATPase activity is expressed as a percentage of the activity in control incubations where protein kinases were not stimulated. Other information as in Fig. 4. *Significantly different from the corresponding incubation with zero calcium concentration; Student's t-test, P<0.01.

View larger version (18K): [in a new window]   Fig. 7. Arrhenius plots showing Na+/K+-ATPase maximal activity as a function of temperature (Kelvin) for (A) foot muscle and (B) hepatopancreas from O. lactea. Values are means ± s.e.m., N=4 independent preparations of enzyme. Assay mixtures were pre-incubated for 5 min in water baths set to specified temperatures prior to initiating reactions by the addition of Mg2+.ATP. Assays were conducted at maximal substrate concentrations. Arrhenius activation energy (EA) was calculated from the linear part of the graphs.

View larger version (17K): [in a new window]   Fig. 8. Western blots showing Na+/K+-ATPase protein content in (A) foot muscle and (B) hepatopancreas of active and 10-day estivated snails. Representative blots are shown for four independent samples. Band densities were normalized against the combined measured densities of three protein bands stained strongly with Coomassie Blue that did not change between control and estivated states, and histograms show the mean normalized band densities (± s.e.m., N=10 for foot muscle, N=8 for hepatopancreas). The band shown corresponds to 100 kDa, as determined from a standard curve developed using Kaleidoscope protein size markers, and is consistent with the expected alpha subunit size of Na+/K+-ATPase.

View larger version (22K): [in a new window]   Fig. 9. Structural unfolding and enzymatic inhibition by urea of foot muscle Na+/K+-ATPase from active and estivating snails. Enzyme extracts were denatured overnight with different concentrations of urea and then either treated with thermolysin or assayed under optimal conditions. (A) Mean relative Na+/K+-ATPase protein remaining (as quantified from band intensities on western blots) after overnight incubation with urea and subsequent proteolysis with thermolysin; values are means ± s.e.m., N=3 independent trials. (B) Mean relative activity of foot muscle Na+/K+-ATPase from active and estivating snails as a function of overnight incubation with different concentrations of urea; values are means ± s.e.m., N=3.