环磷酸腺苷依赖性蛋白激酶催化的合成肽中羟脯氨酸的磷酸化作用。

Phosphorylation of hydroxyproline in a synthetic peptide catalyzed by cyclic AMP-dependent protein kinase.

作者信息

Feramisco J R, Kemp B E, Krebs E G

出版信息

J Biol Chem. 1979 Aug 10;254(15):6987-90.

Abstract

The cyclic AMP-dependent protein kinase catalyzes the phosphorylation of hydroxyproline present in the heptapeptide, Leu-Arg-Arg-Ala-Hyp-Leu-Gly. The Km value for the reaction with this substrate was high (approximately 18 mM) compared to the Km values reported for the analogous threonine and serine-containing peptides, which were 0.59 mM and 0.016 mM, respectively (Kemp, B.E., Graves, D.J., Benjamini, E., and Krebs, E.G. (1977) J. Biol. Chem. 252, 4888-4894). The Vmax value with the hydroxyproline-containing peptide was 1 mumol . min-1 mg-1 in contrast to Vmax values of 6 mumol . min-1 mg-1 and 20 mumol . min-1 mg-1 for the threonine- and serine-containing peptides, respectively. Phosphate esterified to hydroxyproline present in the peptide was relatively stable in hot alkali, only 10% being released as Pi within 30 min in 0.1 N NaOH at 100 degrees C, whereas all of the phosphate was released from the phosphoserine peptide analogue under these conditions. Phosphohydroxyproline in the peptide was also more stable to acid (5.7 N HCl, 110 degrees C) than phosphoserine, the time for 50% release as Pi being 15 h in contrast to 6 h for the latter.

摘要

环磷酸腺苷依赖性蛋白激酶催化七肽Leu-Arg-Arg-Ala-Hyp-Leu-Gly中存在的羟脯氨酸的磷酸化。与报道的含苏氨酸和丝氨酸的类似肽的Km值相比，与该底物反应的Km值较高（约18 mM），含苏氨酸和丝氨酸的类似肽的Km值分别为0.59 mM和0.016 mM（肯普，B.E.，格雷夫斯，D.J.，本杰明尼，E.，和克雷布斯，E.G.（1977年）《生物化学杂志》252，4888 - 4894）。含羟脯氨酸肽的Vmax值为1 μmol·min⁻¹·mg⁻¹，而含苏氨酸和丝氨酸肽的Vmax值分别为6 μmol·min⁻¹·mg⁻¹和20 μmol·min⁻¹·mg⁻¹。肽中与羟脯氨酸酯化的磷酸在热碱中相对稳定，在100℃下于0.1 N NaOH中30分钟内只有10%以无机磷酸（Pi）形式释放，而在这些条件下所有磷酸都从磷酸丝氨酸肽类似物中释放出来。肽中的磷酸羟脯氨酸对酸（5.7 N HCl，110℃）也比磷酸丝氨酸更稳定，50%以Pi形式释放的时间为15小时，而后者为6小时。