LIVER STEROL CARRIER PROTEINS--EFFECTS OF ETHANOL

肝脏甾醇载体蛋白——乙醇的影响

基本信息

批准号：
2748451
负责人：
WELLINGTON GIBSON WOOD
金额：
$ 16.54万
依托单位：
UNIVERSITY OF MINNESOTA
依托单位国家：
美国
项目类别：
财政年份：
1996
资助国家：
美国
起止时间：
1996-08-01 至 2001-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2748451
关键词：
alcoholic beverage consumption binding proteins chemical binding cholesterol ethanol fatty acid binding protein fluorescent dye /probe intermolecular interaction laboratory mouse lipid transport liver metabolism membrane permeability nuclear magnetic resonance spectroscopy phospholipids protein structure function steroid metabolism transport proteins

项目摘要

DESCRIPTION (from Applicant's Abstract): Both lipids and proteins are sites of ethanol action. There have been an extensive number of studies on effects of ethanol on lipid structure. However, while quite a few studies examined effects of ethanol on protein function, there is little information known on how ethanol actually alters protein activity. It has been previously proposed that ethanol directly binds to proteins, but that conclusion has been largely based on effects of ethanol o protein function. Another issue is if ethanol directly binds to a protein what are the consequences of that binding on protein function. Preliminary data indicate that ethanol indeed directly binds to certain lipid transfer proteins, i.e., sterol carrier binding protein-2 (SCP-2), liver-fatty acid binding protein (L-FABP) and bovine serum albumin (BSA). In addition, we observed that ethanol displaced cis-parinaric acid from some, but not all fatty acid-binding sites on BSA. Thus, not only does ethanol directly bind to some proteins but it selectively binds to certain hydrophobic sites. It is the overall hypothesis of this new application that ethanol directly binds to hydrophobic areas of SCP-2 and L-FABP, two proteins that are affected by chronic ethanol consumption. We further proposed that ethanol inhibits the binding of endogenous ligands to SCP-2 and L-FABP and modifies sterol transport between membranes. These hypotheses will be examined using the powerful combination of fluorescence spectroscopy and 13C NMR relaxation technique. The specific aims of this application are: 1) characteristics of cholesterol phospholipids and fatty acids binding to SCP-2 and L-FABP will be examined in the absence and presence of ethanol in vitro using fluorescence techniques; 2) the binding geometry of ethanol and fatty acids to SCP-2 and L-FABP will be studied using 13C- ethanol NMR relaxation data; and 3) effects of SCP-2 and L-FABP enhanced sterol exchange between liver membranes will be evaluated. Chronic alcohol abuse is the most important cause of morbidity and mortality from liver disease in the United States. Approximately 90 percent of heavy alcohol drinkers develop fatty liver, resulting mainly from marked abnormalities in hepatic lipid metabolism, specifically, from accumulation of triacylglycerols and other lipids. In addition to lipids accumulating in liver with chronic ethanol consumption, proteins accumulate as well. Cytosolic proteins are largely responsible for this increase and L-FABP which binds fatty acids and cholesterol accounted for 22 percent of the total increase in liver cytosolic proteins in chronic ethanol treated rats. Therefore, it is important to understand how ethanol may directly act on L- FABP and SCP-2 with respect to binding, displacement of endogenous ligands, and lipid transport.

描述（来自申请人的摘要）：脂质和蛋白质都是乙醇作用的位置。有很多研究对乙醇对脂质结构的影响。但是，虽然很多研究检查了乙醇对蛋白质功能的影响，几乎没有有关乙醇如何实际改变蛋白活性的信息。它以前已提出乙醇直接与蛋白质结合，但是，这一结论主要基于乙醇的影响蛋白质功能。另一个问题是乙醇是否直接与蛋白质对蛋白质功能的结合带来了什么后果。初步数据表明乙醇确实直接与某些脂质转移蛋白，即固醇载体结合蛋白2（SCP-2），肝脂肪酸结合蛋白（L-FABP）和牛血清白蛋白（BSA）。此外，我们观察到乙醇置换了顺式 - 果皮来自BSA上的某些但不是所有脂肪酸结合位点的酸。因此，不是乙醇直接与某些蛋白质结合，但有选择地与某些疏水部位结合。这是总体假设乙醇直接与疏水区域结合的新应用 SCP-2和L-FABP，两种受慢性乙醇影响的蛋白质消耗。我们进一步提出乙醇抑制了内源配体至SCP-2和L-FABP并修饰固醇转运在膜之间。这些假设将使用强大的荧光光谱和13C NMR松弛的组合技术。该应用程序的具体目的是：1）胆固醇磷脂和与脂肪酸结合的特征在不存在和存在乙醇的情况下，将检查SCP-2和L-FABP 使用荧光技术在体外； 2）结合几何形状将使用13C-将研究乙醇和脂肪酸至SCP-2和L-FABP 乙醇NMR松弛数据； 3）SCP-2和L-FABP增强的影响将评估肝膜之间的固醇交换。慢性的酗酒是发病和死亡率的最重要原因来自美国的肝病。大约90％大量酒精饮用者患脂肪肝，主要由标记肝脂质代谢异常，特别是从三酰甘油和其他脂质的积累。此外慢性乙醇消耗，蛋白质积聚在肝脏中的脂质也积累。胞质蛋白在很大程度上为此负责增加和L-FABP结合脂肪酸和胆固醇对于肝胞质蛋白总升高的22％慢性乙醇治疗的大鼠。因此，重要的是要了解乙醇如何直接在L-Fabp和SCP-2上起作用结合，内源配体的位移和脂质转运。