Targeting of the catalytic subunit of protein phosphatase-1 to the glycolytic enzyme phosphofructokinase.
In this study, we demonstrate that the catalytic subunit of rabbit muscle protein phosphatase-1 (PP1) binds to muscle phosphofructokinase (6-phosphofructo-1-kinase, PFK). A protein of 85 kDa was isolated from rat muscle by affinity chromatography on PP1-Sepharose and was identified as phosphofructokinase by partial amino acid sequence analysis. This novel finding of a protein-protein interaction between PP1 and PFK was confirmed by reciprocal experiments in which the binding of PP1 to PFK-agarose was demonstrated. Elution of PP1 from PFK-agarose was maximal at ca. 0.4 M NaCl. The specificity of binding was demonstrated by isolation of PP1 from a partially purified rabbit muscle PP1 preparation. All four known isoforms of PP1 (PP1alpha, PP1gamma1, PP1gamma2, and PP1delta) were shown to bind to PFK-agarose. The activity of PP1 was only partially inhibited by PFK. The preformed complex between PP1 and PFK did not bind to inhibitor-2-Sepharose. The stoichiometry of binding of PP1 to the PFK monomer was found to be 1:1 in the isolated PP1.PFK complex. An interaction between PP1 and PFK in muscle extracts was demonstrated by their coimmunoprecipitation. Our findings raise the interesting possibility that PP1 may be targeted to PFK, and may be physiologically relevant in the context that PFK and other glycolytic enzymes have been shown to be micro-compartmentalized by binding to F-actin. This in turn points to a role for PP1 in control of glycolytic flux by protein phosphorylation-dephosphorylation mechanisms.
Zhao
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Department of Biochemistry and Molecular Biology, University of Miami School of Medicine, Miami, Florida 33101, USA.
Lee
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D000595:Amino Acid Sequence; D000818:Animals; D002384:Catalysis; D002846:Chromatography, Affinity; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D006019:Glycolysis; D007163:Immunosorbent Techniques; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018482:Muscle, Skeletal / Q000201:enzymology; D010732:Phosphofructokinase-1 / Q000737:chemistry / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D054645:Protein Phosphatase 1; D011817:Rabbits; D051381:Rats
D007527:Isoenzymes; D010732:Phosphofructokinase-1; D017868:Cyclic AMP-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1