Immunological mapping of fine molecular surface structures of citrate synthase enzymes from different cell types.
Immunological mapping of fine molecular surface structures of citrate synthase enzymes from different cell types.
复制标题
来自不同细胞类型的柠檬酸合酶的精细分子表面结构的免疫学图谱。
DOI：
10.1002/jmr.300040206
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发表时间：
1991
期刊：
影响因子：
--
通讯作者：
Srere,PA
中科院分区：
文献类型：
--
作者：
Nemeth,P;Small,WC;Evans,CT;Zhi,W;Persson,LO;Srere,PA
柠檬酸合成酶(EC 4.1.3.7)作为有氧能量代谢的关键酶存在于所有生物体内，是已知的最具系统发育保守性的酶之一。然而，在体外稳定性、对pH变化的耐受性、自聚合度等其他参数方面，不同来源的柠檬酸合成明显不同。即使在同一物种内酶的不同亚型之间也可以观察到这些差异。记录这些差异的数据表明，三级结构可能会发生高度差异。因此，本研究利用抗猪柠檬酸合成酶(α-PCs)和酵母柠檬酸合成酶-2(α-YCS-2)的单抗家族，用免疫学方法研究了酵母、猪、大鼠、番茄和大肠杆菌中柠檬酸合成酶的表面特征。来自酵母、番茄、猪和大鼠细胞的线粒体柠檬酸合成酶的酶表位具有高度的同源性。主要的差异是发现起源于Frome的六聚体柠檬酸合成酶。与从真核细胞制备的二聚体形式进行了比较。在高度同源的过氧化体和线粒体酵母柠檬酸盐合成之间只检测到适度的相似之处。此外，线粒体柠檬酸合成酶(CS-1)的一个催化残基(重组猪上的H274R和酵母酶上的H313R)的点突变导致与过氧化体同工酶(CS-2)的免疫学相似性显著增加。根据CS-2在酵母中进化的可能机制，对这些发现进行了讨论。
Citrate synthase (EC 4.1.3.7), which is present in all living organisms as a key enzyme in aerobic energy metabolism, is one of the most highly phylogenetically conserved enzymes known in terms of its primary and active site structure. However, in terms of other parameters such asin vitrostability, tolerance to changes in pH, degree of self‐polymerization, etc., citrate syntheses different sources are markedly different. These divergences can be observed even between isoforms of the enzyme within the same species. Data documenting these diversities suggest that a high degree of difference in tertiary structures may occur. Therefore, the surface profiles of citrate synthase enzymes from yeast, pig, rat, tomato andEscherichia coliwere investigated with immunological methods using monoclonal antibody families generated against either pig citrate synthase (α‐PCS) or yeast citrate synthase‐2 (α‐YCS‐2). A high degree of homology of enzyme epitopes was detected on the mitochondrial citrate synthases originating from yeast, tomato, pig and rat cells. Major differences were found between the hexameric citrate synthase originating fromE. colicompared with those dimeric forms prepared from eukaryotic cells. Only modest similarities were detected between the highly homologous peroxisomal and mitochondrial yeast citrate syntheses. Furthermore, a point mutation of one of the catalytic residues (H274R on recombinant pig and H313R on yeast enzyme) of mitochondrial citrate synthase (CS‐1) resulted in a significant increase in immunological similarity with the peroxisomal isoenzyme (CS‐2). These findings are discussed in terms of the possible mechanism of evolution of CS‐2 in yeast.