Background. Pyruvate kinase (PK) is responsible for the final reaction in glycolysis. As PK is a glycolytic control point, the analysis of PK posttranslational modifications (PTM) and kinetic changes reveals a key piece of the reorganization of energy metabolism in an anoxia tolerant vertebrate. Methods. To explore PK regulation, the enzyme was isolated from red skeletal muscle and liver of aerobic and 20-hr anoxia-exposed red eared-slider turtles (Trachemys scripta elegans). Kinetic analysis and immunoblotting were used to assess enzyme function and the corresponding covalent modifications to the enzymes structure during anoxia. Results. Both muscle and liver isoforms showed decreased affinity for phospho- enolpyruvate substrate during anoxia, and muscle PK also had a lower affinity for ADP. I50 values for the inhibitors ATP and lactate were lower for PK from both tissues after anoxic exposure while I50 L-alanine was only reduced in the liver. Both isozymes showed significant increases in threonine phosphorylation (by 42% in muscle and 60% in liver) and lysine methylation (by 43% in muscle and 70% in liver) during anoxia which have been linked to suppression of PK activity in other organisms. Liver PK also showed a 26% decrease in tyrosine phosphorylation under anoxia. Discussion. Anoxia responsive changes in turtle muscle and liver PK coordinate with an overall reduced activity state. This reduced affinity for the forward glycolytic reaction is likely a key component of the overall metabolic rate depression that supports long term survival in anoxia tolerant turtles. The coinciding methyl- and phospho- PTM alterations present the mechanism for tissue specific enzyme modification during anoxia.

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Keywords Anoxia, Enzyme regulation, Glycolysis, Posttranslational modifications, Pyruvate kinase, Trachemys scripta elegans
Persistent URL
Journal PeerJ
Mattice, A.M. (Amanda M.S.), MacLean, I.A. (Isabelle A.), Childers, C.L. (Christine L.), & Storey, K. (2018). Characterization of pyruvate kinase from the anoxia tolerant turtle, Trachemys scripta elegans: A potential role for enzyme methylation during metabolic rate depression. PeerJ, 2018(6). doi:10.7717/peerj.4918