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Cryobiology. 2017 Aug;77:25-33. doi: 10.1016/j.cryobiol.2017.06.002. Epub 2017 Jun 16.

Regulation of pyruvate kinase in skeletal muscle of the freeze tolerant wood frog, Rana sylvatica.

Author information

1
Institute of Biochemistry, Department of Biology, Carleton University, 1125 Colonel By Drive, Ottawa, Ontario K1S 5B6, Canada.
2
Institute of Biochemistry, Department of Biology, Carleton University, 1125 Colonel By Drive, Ottawa, Ontario K1S 5B6, Canada. Electronic address: kenneth.storey@carleton.ca.

Abstract

The wood frog (Rana sylvatica) can survive the winter in a frozen state, in which the frog's tissues are also exposed to dehydration, ischemia, and anoxia. Critical to wood frog survival under these conditions is a global metabolic rate depression, the accumulation of glucose as a cryoprotectant, and a reliance on anaerobic glycolysis for energy production. Pyruvate kinase (PK) catalyzes the final reaction of aerobic glycolysis, generating pyruvate and ATP from phosphoenolpyruvate (PEP) and ADP. This study investigated the effect of each stress condition experienced by R. sylvatica during freezing, including dehydration and anoxia, on PK regulation. PK from muscle of frozen and dehydrated frogs exhibited a lower affinity for PEP (Km = 0.098 ± 0.003 and Km = 0.092 ± 0.008) than PK from control and anoxic conditions (Km = 0.065 ± 0.003 and Km = 0.073 ± 0.002). Immunoblotting showed greater serine phosphorylation on muscle PK from frozen and dehydrated frogs relative to control and anoxic states, suggesting a reversible phosphorylation regulatory mechanism for PK activity during freezing stress. Furthermore, PK from frozen animals exhibited greater stability under thermal and urea-induced denaturing conditions than PK from control animals. Phosphorylation of PK during freezing may contribute to mediating energy conservation and maintaining intracellular cryoprotectant levels, as well as increase enzyme stability during stress.

KEYWORDS:

Freeze tolerance; Metabolic rate depression; Pyruvate kinase; Rana sylvatica

PMID:
28600082
DOI:
10.1016/j.cryobiol.2017.06.002
[Indexed for MEDLINE]

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