Wetterau J R, Aggerbeck L P, Laplaud P M, McLean L R
Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, Ohio 45267-0575.
Biochemistry. 1991 May 7;30(18):4406-12. doi: 10.1021/bi00232a006.
The microsomal triglyceride-transfer protein (MTP), which catalyzes the transport of triglyceride and cholesteryl ester between membranes, is a complex composed of two proteins having apparent molecular weights of 58,000 and 88,000. The 58,000 molecular weight component of MTP has been identified as the multifunctional protein, protein disulfide isomerase (PDI). The multisubunit nature of MTP as well as the presence of PDI as one of the subunits distinguishes this protein from previously characterized lipid-transfer proteins. In this study, we have more clearly defined structural elements of MTP that may play important functional roles. The molecular weight of the transfer protein complex was determined to be 150,000 by sedimentation equilibrium experiments performed at three different speeds, suggesting that MTP is a complex of one PDI and one 88,000 molecular weight polypeptide (88K). Following SDS-polyacrylamide gel electrophoresis, the Coomassie Blue staining intensity of PDI in a known amount of MTP was compared to that of known amounts of a PDI standard. A 1 to 0.98-1.30 ratio of PDI to 88K was determined, confirming the 1:1 stoichiometry of MTP. The sedimentation coefficient (5.85) determined by analytical ultracentrifugation and the Stokes radius (47 A) determined by polyacrylamide gradient gel electrophoresis indicate that the 150,000 molecular weight MTP complex is asymmetric and/or has an unusually high water of hydration. PDI and 88K form a stable protein complex; there was no evidence of a dissociation-reassociation reaction occurring between the two components. Analysis of far-ultraviolet circular dichroism spectra revealed MTP has about 28% alpha-helical and 28% beta-structural content.(ABSTRACT TRUNCATED AT 250 WORDS)
微粒体甘油三酯转运蛋白（MTP）催化甘油三酯和胆固醇酯在膜之间的转运，它是一种由两种蛋白质组成的复合物，这两种蛋白质的表观分子量分别为58,000和88,000。MTP的58,000分子量组分已被鉴定为多功能蛋白——蛋白二硫键异构酶（PDI）。MTP的多亚基性质以及作为亚基之一的PDI的存在，使该蛋白有别于先前表征的脂质转运蛋白。在本研究中，我们更明确地界定了MTP中可能发挥重要功能作用的结构元件。通过在三种不同速度下进行沉降平衡实验，确定转运蛋白复合物的分子量为150,000，这表明MTP是一个由一个PDI和一个88,000分子量多肽（88K）组成的复合物。在十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳后，将已知量的MTP中PDI的考马斯亮蓝染色强度与已知量的PDI标准品的染色强度进行比较。确定PDI与88K的比例为1比0.98 - 1.30，证实了MTP的1:1化学计量比。通过分析超速离心法测定的沉降系数（5.85）和通过聚丙烯酰胺梯度凝胶电泳测定的斯托克斯半径（47 Å）表明，150,000分子量的MTP复合物是不对称的和/或具有异常高的水化水。PDI和88K形成稳定的蛋白质复合物；没有证据表明这两种组分之间发生解离 - 重新结合反应。远紫外圆二色光谱分析表明，MTP含有约28%的α - 螺旋和28%的β - 结构成分。（摘要截短于250字）
