Phosphorylation of rat hepatic fructose-1,6-bisphosphatase and pyruvate kinase.

Fructose-1,6-bisphosphatase from rat liver was phosphorylated with cyclic AMP-dependent protein kinase and [gamma-32P]ATP. Brief exposure of the 32P-labeled enzyme to trypsin removed all radioactivity from the enzyme core and produced a single-labeled peptide. The partial sequence of the 17-amino acid peptide was found to be Ser-Arg-Pro-Ser(P)-Leu-Pro-Leu-Pro-(Ser2, Glx2, Pro2, Leu, Arg2). The kinetics of cyclic AMP-dependent protein kinase-catalyzed phosphorylation of native fructose bisphosphatase were compared with those of rat liver type L pyruvate kinase where the sequence around the phosphoserine is known (Arg-Arg-Ala-Ser(P)-Val; Hjelmquist, G., Anderson, J., Edlund, B., and Engstrom, L. (1974) Biochem. Biophys. Res. Commun. 61, 559-563). The Km for pyruvate kinase (17 microM) was less than that for fructose bisphosphatase (58 microM); the Vmax was about 3-fold greater with pyruvate kinase as substrate. The relationship between the rates of phosphorylation of these native substrates and the amino acid sequences surrounding the phosphorylated sites is discussed.