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Cell Biology Poster Session






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Protein Kinase A from Bat Skeletal Muscle: A Kinetic Study of the Enzyme from a Hibernating Mammal

Holden, C.P. (Department of Pharmacology and Therapeutics, University of Manitoba, Canada)
Storey, J. (Departments of Chemistry and Biology, Carleton University, Canada)
Storey, K.B. (Departments of Chemistry and Biology, Carleton University, Canada)

Contact Person: Clark P. Holden (cholden@cc.umanitoba.ca)


Abstract

The catalytic subunit of adenosine 3'-5'-cyclic monophosphate-dependent protein kinase (PKAc) was purified to homogeneity from skeletal muscle of the little brown bat, Myotis lucifugus. Final specific activity was 205 nmol phosphate transferred/min/mg protein at 22C. Identification of the enzyme as a protein kinase A was confirmed through the use of specific PKA inhibitors. PKAc molecular weight was 54.6 3.5 kDa. Km values for Kemptide and Mg-ATP were 9.1 0.2 micromolar and 94.1 4.5 micromolar at 37C, respectively; both values decreased significantly at 5C. Neutral salts had little effect on enzyme activity (I50 values > 400 mM), but NaF did with an I50 value of 38 mM. Arrhenius plots showed evidence of a temperature-dependent conformational change in PKAc. Calculated activation energies were 5.6 0.46 kJ/mol at temperatures above 10C and 29.5 2.0 kJ/mol below. The pH optimum of bat PKAc also changed dramatically with temperature falling from 8.5 at 37C to 5.5 at 5C, an effect that could substantially limit enzyme activity in vivo at the low body temperatures seen during hibernation. Overall, low temperature had both positive (increased %PKAc, reduced Km values, increased I50 values for salts) and negative (increased activation energy, acidic shift of pH optimum) effects on PKAc. However, the substantial positive effects of low temperature on the enzyme suggest a critical role for continued PKAc action in signal transduction processes in the hibernating bat.

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Poster Number PAholden0436
Keywords: hibernation, Signal, transduction, PKA, skeletal muscle


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Holden, C.P.; Storey, J.; Storey, K.B.; (1998). Protein Kinase A from Bat Skeletal Muscle: A Kinetic Study of the Enzyme from a Hibernating Mammal. Presented at INABIS '98 - 5th Internet World Congress on Biomedical Sciences at McMaster University, Canada, Dec 7-16th. Available at URL http://www.mcmaster.ca/inabis98/cellbio/holden0436/index.html
© 1998 Author(s) Hold Copyright