Hrdý I
Laboratoire de Chimie Physiologique, Université Catholique de Louvain, Belgium.
Folia Parasitol (Praha). 1993;40(3):181-5.
A cytosolic malate dehydrogenase from Tritrichomonas foetus (Riedmüller) was purified to apparent homogeneity by a combination of differential centrifugation, affinity, hydrophobic interaction and ion-exchange chromatography. The purest preparation had specific activity of 309 mumoles.(mg protein)-1.min-1, which corresponded to 50-fold purification. The enzyme representing almost 2% of total cytosolic protein displayed hyperbolic affinity curves for substrate and coenzyme. Km for oxaloacetate and NADH was 17.5 microM and 13 microM, respectively. Subunit size determined by SDS-PAGE and by laser desorption mass spectroscopy was approximately 37 kDa, which corresponds to values reported for most malate dehydrogenases. The native molecular weight determined by gel filtration was 244 kDa, indicating six subunit structure of holoenzyme; this is unusual in comparison with other malate dehydrogenases, which are usually dimers or tetramers.
通过差速离心、亲和、疏水相互作用和离子交换色谱相结合的方法，将胎儿三毛滴虫（Riedmüller）的胞质苹果酸脱氢酶纯化至表观均一。最纯的制剂的比活性为309微摩尔·（毫克蛋白）-1·分钟-1，相当于50倍的纯化倍数。该酶占总胞质蛋白的近2%，对底物和辅酶呈现双曲线亲和力曲线。草酰乙酸和NADH的Km分别为17.5微摩尔和13微摩尔。通过SDS-PAGE和激光解吸质谱法测定的亚基大小约为37 kDa，这与大多数苹果酸脱氢酶报道的值相对应。通过凝胶过滤测定的天然分子量为244 kDa，表明全酶具有六个亚基结构；与其他通常为二聚体或四聚体的苹果酸脱氢酶相比，这是不寻常的。
