Slomiany B L, Murty V L, Slomiany A, Zielenski J, Mandel I D
Biochim Biophys Acta. 1986 Jun 3;882(1):18-28. doi: 10.1016/0304-4165(86)90050-4.
A high molecular weight mucus glycoprotein has been isolated from submandibular saliva of caries-resistant and caries-susceptible individuals by a procedure involving fractionation on Bio-Gel P-100 and A-50 columns followed by equilibrium density-gradient centrifugation in CsCl. The purified caries-resistant mucus glycoprotein displayed a buoyant density of 1.50 and accounted for 9.5% of the dry weight of caries-resistant saliva. The caries-susceptible mucus glycoprotein represented 14.1% of the dry weight of caries-susceptible saliva and gave a buoyant density of 1.43. Both glycoproteins exhibited similar protein and carbohydrate content, but the caries-resistant mucus glycoprotein contained 28.7% less associated lipids and 3-times less covalently bound fatty acids than the caries-susceptible mucus glycoprotein. The associated lipids were represented by neutral lipids, glycolipids and phospholipids, whereas the covalently bound fatty acids consisted mainly of hexadecanoate, octadecanoate and docosanoate. Extraction of associated lipids caused the caries-resistant glycoprotein to band in CsCl gradient at the density of 1.54 and caused the caries-susceptible glycoprotein to band at the density of 1.52. A further shift in the buoyant densities occurred following removal of the covalently bound fatty acids, and both glycoproteins banded at the density of 1.57. While the intact caries-resistant and caries-susceptible glycoproteins were susceptible to proteolysis by pronase, the lipid-rich caries-susceptible glycoprotein was degraded to a lesser extent. Extraction of associated lipids increased the degradation of both glycoproteins, but the caries-susceptible glycoprotein still remained 25% less susceptible. However, the susceptibility to pronase of the delipidated and deacylated caries-resistant and caries-susceptible glycoproteins was essentially identical. The caries-resistant and caries-susceptible mucus glycoproteins also differed in susceptibility to peptic degradation. The apparent Km values for intact caries-resistant and caries-susceptible glycoproteins were 10.5 X 10(-7) M and 8.1 X 10(-7) M, while the values for the delipidated and deacylated caries-resistant and caries-susceptible glycoproteins were 13.0 X 10(-7) M and 12.4 X 10(-7) M. The results suggest that the differences in the content of associated lipids and covalently bound fatty acids are responsible for the different physiochemical characteristics of caries-resistant and caries-susceptible salivary mucus glycoproteins, which may be determining factors in the resistance to caries.
通过包括在Bio - Gel P - 100和A - 50柱上分级分离,随后在CsCl中进行平衡密度梯度离心的方法,从患龋者和易感龋个体的颌下唾液中分离出一种高分子量粘液糖蛋白。纯化后的抗龋粘液糖蛋白浮力密度为1.50,占抗龋唾液干重的9.5%。易感龋粘液糖蛋白占易感龋唾液干重的14.1%,浮力密度为1.43。两种糖蛋白的蛋白质和碳水化合物含量相似,但抗龋粘液糖蛋白所含的结合脂质比易感龋粘液糖蛋白少28.7%,共价结合脂肪酸少3倍。结合脂质由中性脂质、糖脂和磷脂组成,而共价结合脂肪酸主要由十六烷酸、十八烷酸和二十二烷酸组成。提取结合脂质后,抗龋糖蛋白在CsCl梯度中的条带密度变为1.54,易感龋糖蛋白的条带密度变为1.52。去除共价结合脂肪酸后,浮力密度进一步变化,两种糖蛋白的条带密度均为1.57。完整的抗龋和易感龋糖蛋白都易被链霉蛋白酶水解,但富含脂质的易感龋糖蛋白降解程度较小。提取结合脂质增加了两种糖蛋白的降解,但易感龋糖蛋白的降解程度仍低25%。然而,去脂和去酰化的抗龋和易感龋糖蛋白对链霉蛋白酶的敏感性基本相同。抗龋和易感龋粘液糖蛋白对胃蛋白酶降解的敏感性也不同。完整的抗龋和易感龋糖蛋白的表观Km值分别为10.5×10⁻⁷M和8.1×10⁻⁷M,而去脂和去酰化的抗龋和易感龋糖蛋白的值分别为13.0×10⁻⁷M和12.4×10⁻⁷M。结果表明,结合脂质和共价结合脂肪酸含量的差异是抗龋和易感龋唾液粘液糖蛋白不同理化特性的原因,这可能是抗龋能力的决定因素。