STUDIES ON THE ISOLATION AND COMPOSITION OF HUMAN OCULAR MUCIN
STUDIES ON THE ISOLATION AND COMPOSITION OF HUMAN OCULAR MUCIN
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DOI：
10.1016/0014-4835(88)90002-4
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发表时间：
1988-08-01
影响因子：
3.4
通讯作者：
HERP, A
中科院分区：
文献类型：
--
作者：
CHAO, CCW;BUTALA, SM;HERP, A
本文报道了一种从人眼粘液的盐水提取液中分离纯化人眼粘蛋白的方法。通过在Sephadex G-50和Sepharose CL-4 B柱上对来自个体供体粘液池的盐溶性粘液提取物进行连续色谱，实现眼粘蛋白的初步纯化。用碱性β-半乳糖苷酶对一部分这种粘蛋白分离物进行粘蛋白特征性的O-丝氨酰（苏氨酰）-N-乙酰半乳糖胺基键的定量分析。消除和氚化硼氢化物还原。在Bio-Gel P-2过滤后，认为粘蛋白分离物是富含粘蛋白的，该分离物的裂解寡糖含有> 0.5 μ Ci mg-1的氚化半乳糖胺醇，该值代表牛和绵羊颌下参照粘蛋白所观察到的值的至少64%。这些分离物在Sephacryl S-500和DEAE-Trisacryl M柱层析上进行进一步纯化。纯化的粘蛋白具有120 kDa的最小分子量。它由25-30%蛋白质和54-55%碳水化合物组成。其氨基酸和碳水化合物组成具有粘蛋白结构的特征。通过SDS-梯度PAGE验证粘蛋白的纯度。等电聚焦后，在pI 5.0-6.6范围内显示多分散性/微异质性。
A method for the isolation and purification of human ocular mucin from the brief saline extract of human ocular mucus is reported. Initial purification of ocular mucin was achieved by sequential chromatography of the saline-soluble mucus extract from an individual donor''s mucus pool on columns of Sephadex G-50 and Sepharose CL-4B. A portion of such mucin isolate was subjected to quantitative analysis of the O-seryl (threonyl)-N-acetylgalactosaminyl linkage, characteristic of mucins, by alkaline .beta.-elimination and tritiated borohydride reduction. Following Bio-Gel P-2 filtration, the mucin isolate whose cleaved oligosaccharides contained tritiated galactosaminitol > 0.5 .mu.Ci mg-1, a value that represents at least 64% of that observed for bovine and ovine submaxillary reference mucins, was considered to be mucin-rich. These isolates were subjected to further purification on Sephacryl S-500 and DEAE-Trisacryl M column chromatographies. The purified mucin had a minimum molecular weight of 120 kDa. It consisted of 25-30% protein and 54-55% carbohydrate. Its amino acid and carbohydrate compositions are charcteristic of a mucin structure. The purity of the mucin was verified by SDS-gradient PAGE. Upon isoelectric focusing, polydispersity/microheterogeneity were exhibited in the pI range 5.0-6.6.