Robinson G, Chapman B E, Kuchel P W
Eur J Biochem. 1984 Sep 17;143(3):643-9. doi: 10.1111/j.1432-1033.1984.tb08417.x.
The rate of exchange of phosphoryl groups between 2- and 3-phosphoglycerate catalysed by (a) high concentrations (approximately equal to 5.0 mg protein ml-1) of rabbit muscle phosphoglyceromutase and (b) lysed human erythrocytes was measured using saturation and inversion transfer techniques with 31P-NMR spectroscopy. This is the first reported application of these techniques to a study of this particular enzymic reaction either in vitro or in situ in a cell cytosol. Selective irradiation of resonances was achieved by the DANTE pulse sequence which had not previously been used for spin-transfer studies. New equilibrium exchange theory was developed for the simplest model of a two-reactant enzyme-catalysed reaction and this was used to calculate turnover rates for the enzymes. There was a close similarity between the turnover rates calculated from the spin-transfer data obtained from the systems in vitro and in situ and those obtained by conventional enzymic assays, at low enzyme concentrations. This suggested an absence of any homogeneous enzyme-enzyme interactions which modify the kinetics at high protein concentrations either in lysates or in the system in vitro.
利用³¹P - NMR光谱的饱和与反转转移技术，测定了在以下两种情况下，2 - 磷酸甘油酸和3 - 磷酸甘油酸之间磷酰基的交换速率：(a)高浓度（约等于5.0 mg蛋白质/ ml）的兔肌肉磷酸甘油酸变位酶；(b)裂解的人红细胞。这是首次报道将这些技术应用于体外或细胞胞质溶胶中原位研究这种特定酶促反应。通过DANTE脉冲序列实现共振的选择性照射，该序列以前未用于自旋转移研究。针对最简单的双反应物酶促反应模型，开发了新的平衡交换理论，并用于计算酶的周转速率。在低酶浓度下，从体外和原位系统获得的自旋转移数据计算出的周转速率，与通过传统酶促测定获得的周转速率非常相似。这表明在裂解物或体外系统中，不存在任何在高蛋白浓度下改变动力学的均相酶 - 酶相互作用。
