Biochemical Studies on the Shell Gland of Japanese Quail, Coturnix coturnix japonica: 2. Allosteric two Components and their Catalytic Properties of Pyruvate Kinase in the Shell Gland1

Abstract

Pyruvate kinase (EC 2.7.1.40) in Japanese quail shell gland was examined for the multiple components and enzymic properties. Two components were obtained by isoelectric focusing. Isoelectric points of the two components were pH 4.5 (designated SGPK-I) and pH 6.1 (designated SGPK-II), respectively. In the shell gland mucosa, the existence of only SGPK-II was detected. SGPK-I and SGPK-II were eluted from DEAE-cellulose by 0.02M and 0.12M KC1, respectively at pH 7.4 (20mM Tris-HCl). In the absence of fructose-1,6-diphosphate (FDP), SGPK-I and SGPK-II, showed a sigmoidal response with respect to phosphoenolpyruvate (PEP) concentration, while in the presence of FDP, they both exhibited the Michaelis-Menten type response. Kinetical responses of SGPK-I and SGPK-II were markedly influenced by pH, K+ and Mg2+ concentration even in the presence of FDP. No significant difference in apparent Km value for PEP of the two components was obtained. The inhibition of the two components by ATP was pH-dependent and was reversed by the presence of FDP. The two components from the shell gland were inhibited by Ca2+. The inhibition by Ca2+ was dependent on pH, K+, and Mg2+ concentration. FDP altered the Km value for PEP several folds smaller, of the two components inhibited by Ca2+, but did not change the Vmax. During the egg formation process, the ratio of the summed activity of SGPK-I to that of SGPK-II varied from 0.5 (minimum) to 2.0 (maximum), depending on the position of a developing egg in the oviduct. Physiological significance of the two components in the shell gland is discussed.