Cyclic nucleotide phosphodiesterase and 5′-nucleotidase: A coupled system

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Abstract
Evidence is presented that multiple forms of cyclic nucleotide phophodiesterase (PDE) activity chromatographically separated from the soluble fraction of bovine hypothalamus are co-eluted with multiple forms of 5′-nucleotidase (5′N) activity. The enzymes could not be resolved from each other by anion-exchange chromatography on DEAE-TSK; by affinity chromatography on phenyl-, blue-, concanavalin A-, 5′ AMP-sepharose, cAMP-silica gel; or by gel filtration on sephacryl S-200. The catalytic activities were found to be associated with the tetrameric, dimeric, and monomeric forms of the enzymes. The molecular weights determined by gel filtration or by SDS-gel electrophoresis were 220, 114, and 57 kDa, respectively. Kinetic analysis revealed that the first-order rate constant for 5′ AMP hydrolysis measured in the reactions: cAMP→5′AMP→adenosine was 100 times higher than that in the reaction: 5′AMP→adenosine. Thus, functional interrelation between PDE and 5′N was expressed in drastic acceleration of the consecutive reactions: cAMP →5′AMP→adenosine. The results confirm the conclusion about the existence of a multienzyme system involving PDE and 5′N or of a single bifunctional enzyme in brain tissue.