Bansal M P, Cook R G, Danielson K G, Medina D
Department of Cell Biology, Baylor College of Medicine, Houston, Texas.
J Biol Chem. 1989 Aug 15;264(23):13780-4.
In a previous study, we purified three selenium-binding proteins (molecular masses 56, 14, and 12 kDa) from mouse liver using column chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The aim of the present study was to determine the amino acid sequence of the 14-kDa protein thereby establishing any relationship with known proteins. Although the amino terminus of the 14-kDa protein was blocked, separate in situ digestions of the protein with endoproteinases Glu-c and Lys-c gave overlapping peptides that provided a continuous sequence of 93 amino acids. This sequence exhibited a 92.5% sequence homology with rat liver fatty acid-binding protein. In situ enzymatic digestion and partial sequencing of a 12-kDa selenium-binding protein revealed identical homology to the 14-kDa protein. The 14-kDa protein bound specifically to an oleate-affinity column from which the protein and 75Se coeluted. Delipidation or sodium dodecyl sulfate treatment failed to remove 75Se from the protein, indicating that the selenium moiety was tightly bound to the protein. These observations confirm that the mouse liver selenium-binding 14-kDa protein is a fatty acid-binding protein. The nature of the selenium linkage to the protein still needs to be explored.
在之前的一项研究中，我们使用柱色谱法和十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳从小鼠肝脏中纯化出了三种硒结合蛋白（分子量分别为56、14和12 kDa）。本研究的目的是确定14 kDa蛋白的氨基酸序列，从而确定其与已知蛋白的关系。尽管14 kDa蛋白的氨基末端被封闭，但用内肽酶Glu-c和Lys-c对该蛋白进行原位消化分别得到了重叠肽段，这些肽段提供了一个由93个氨基酸组成的连续序列。该序列与大鼠肝脏脂肪酸结合蛋白具有92.5%的序列同源性。对12 kDa硒结合蛋白进行原位酶切和部分测序，发现其与14 kDa蛋白具有相同的同源性。14 kDa蛋白特异性结合到油酸亲和柱上，该蛋白与75Se从柱上共同洗脱。脱脂或十二烷基硫酸钠处理均未能从该蛋白中去除75Se，这表明硒部分与该蛋白紧密结合。这些观察结果证实，小鼠肝脏中14 kDa的硒结合蛋白是一种脂肪酸结合蛋白。硒与该蛋白的连接性质仍有待探索。
