Takagi A, Slomiany B L, Kosmala M, Slomiany A
Biochim Biophys Acta. 1986 Oct 29;884(1):1-10. doi: 10.1016/0304-4165(86)90219-9.
The resistance to proteolysis by pepsin of gastric mucus glycoprotein synthesized by tissue culture in the presence and absence of 0.1 M ethanol was investigated. The glycoprotein product of ethanol-supplemented culture was found to contain 68% less associated lipids and 81% less covalently bound fatty acids, but exhibited unaltered content of carbohydrate and protein. The lipid and fatty acyl deficient glycoprotein was 5-times more rapidly and 2-3-times more extensively degraded by pepsin than the glycoprotein synthesized in the absence of ethanol. Following delipidation with organic solvents and deacylation with hydroxylamine both glycoproteins were digested at the same rate and degraded to the same extent. The lower content of fatty acyl residues markedly affected the overall pattern of the proteolytic fragments identified by SDS gel electrophoresis. The peptides corresponding to the acylated fragments of control were degraded and an increase in the amount of smaller peptides was observed. The in vitro assays of the fatty acyltransferase activity towards the substrates obtained from control and alcohol-containing cultures revealed that the enzyme activity was similar and increased proportionally with increased concentration of both glycoprotein substrates and enzyme. However, addition of 0.1 M ethanol to the assay tubes containing complete incubation mixture decreased the acylation of either glycoprotein by 40%. Based on the results presented here, and on previous studies of mucus glycoprotein synthesis in the presence of ethanol, we conclude that ethanol interferes with the process of acylation of mucus glycoprotein with fatty acids.
研究了在有和没有0.1M乙醇存在的情况下，通过组织培养合成的胃黏液糖蛋白对胃蛋白酶蛋白水解的抗性。发现补充乙醇培养的糖蛋白产物所含的结合脂质减少68%，共价结合脂肪酸减少81%，但碳水化合物和蛋白质含量未改变。与在没有乙醇的情况下合成的糖蛋白相比，缺乏脂质和脂肪酰基的糖蛋白被胃蛋白酶降解的速度快5倍，降解程度高2至3倍。用有机溶剂脱脂并用羟胺脱酰后，两种糖蛋白以相同的速率被消化并降解到相同的程度。脂肪酰基残基含量较低显著影响了通过SDS凝胶电泳鉴定的蛋白水解片段的总体模式。与对照的酰化片段相对应的肽被降解，并且观察到较小肽的量增加。对从对照和含酒精培养物中获得的底物的脂肪酰基转移酶活性的体外测定表明，酶活性相似，并且随着糖蛋白底物和酶浓度的增加成比例增加。然而，向含有完全孵育混合物的测定管中加入0.1M乙醇会使任一种糖蛋白的酰化作用降低40%。基于此处给出的结果以及先前在有乙醇存在的情况下对黏液糖蛋白合成的研究，我们得出结论，乙醇会干扰黏液糖蛋白与脂肪酸的酰化过程。
