Asthir B, Singh R
Department of Biochemistry, Punjab Agricultural University, Ludhiana, India.
Indian J Biochem Biophys. 1997 Dec;34(6):529-34.
Neutral invertase from nodules of chickpea (Cicer arietinum L.) was isolated and purified by ammonium sulphate fractionation, gel filtration and DEAE-cellulose column chromatography. The purified enzyme was stable between 0 to 40 degrees C beyond which it was irreversibly denatured. Optimum temperature and pH of the enzyme were 37 degrees C and 7.0, respectively. K(m) for sucrose was 14.2 mM and Vmax was 4.8 mumole hr-1. The enzyme was inhibited by several metal ions. From the temperature effect on K(m) and Vmax values, the energy of activation (Ea), enthalpy change (delta H) and entropy change (delta S) of the enzyme were calculated to be 147 kJmol-1, -4.10 kJmol-1 and -2.33 JK-1mol-1, respectively. By employing photo-oxidation and chemical modification and by studying the effect of pH on K(m) and Vmax, the involvement of sulphydryl-, imidazole- and alpha-amino groups in the active site of the enzyme has been indicated.
通过硫酸铵分级沉淀、凝胶过滤和DEAE - 纤维素柱色谱法从鹰嘴豆（Cicer arietinum L.）根瘤中分离并纯化了中性转化酶。纯化后的酶在0至40摄氏度之间稳定，超过此温度会发生不可逆变性。该酶的最适温度和pH分别为37摄氏度和7.0。蔗糖的米氏常数（K(m)）为14.2 mM，最大反应速度（Vmax）为4.8微摩尔·小时-1。该酶受到几种金属离子的抑制。根据温度对K(m)和Vmax值的影响，计算出该酶的活化能（Ea）、焓变（delta H）和熵变（delta S）分别为147 kJmol-1、-4.10 kJmol-1和-2.33 JK-1mol-1。通过光氧化和化学修饰以及研究pH对K(m)和Vmax的影响，表明了巯基、咪唑基和α - 氨基在酶活性位点中的作用。
