Miller L J, Springer T A
J Immunol. 1987 Aug 1;139(3):842-7.
The p150,95 cell surface protein is a member of a family of heterodimeric leukocyte adhesion proteins that have homologous alpha subunits, each noncovalently associated with a common beta subunit. In this report we have metabolically labeled the U937 cell line at various timepoints during its phorbol myristic acetate-induced maturation to examine the kinetics of synthesis of these proteins during monocytic differentiation, and their maturation and glycosylation. The p150,95 alpha subunit was immunoprecipitated with p150,95-specific monoclonal antibody (MAb), or an antiserum to the denatured, purified alpha X subunit. The glycosylation and polypeptide chain length of the p150,95, Mac-1, and lymphocyte function associated antigen (LFA-1) alpha and beta subunits were compared by immunoprecipitation with subunit specific MAb and antisera, and by digestion with Endo H and N-glycanase. The p150,95 alpha subunit is synthesized as a precursor of 146,000 Mr, has five to six N-linked oligosaccharides, and has a polypeptide chain backbone of 132,000 Mr. Over 50% of the carbohydrate on the mature alpha subunit of 150,000 Mr was sensitive to Endo H digestion. The p150,95 alpha and beta precursors can associate before maturation into the mature form. Conversion to the mature form was accompanied by loss of reactivity with the antiserum to the denatured alpha X subunit, suggesting a change in conformation. Mac-1 and LFA-1 alpha subunits have precursors of 160,000 Mr and 165,000 Mr, respectively, and contain N-linked carbohydrates. The polypeptide chain length for the Mac-1 alpha subunit is 137,000 Mr, and for LFA-1 is 149,000 Mr. Only 14% of the oligosaccharide on the mature LFA-1 alpha subunit was sensitive to Endo H, suggesting that unlike p150,95, most is converted to the complex type. The differences noted in the Mr of the three homologous alpha subunits are therefore due to differences in both polypeptide chain length and carbohydrate processing during biosynthesis.
p150,95细胞表面蛋白是异二聚体白细胞粘附蛋白家族的成员，该家族具有同源的α亚基，每个α亚基都与一个共同的β亚基非共价结合。在本报告中，我们在佛波醇肉豆蔻酸酯诱导的U937细胞系成熟过程中的不同时间点对其进行代谢标记，以研究单核细胞分化过程中这些蛋白的合成动力学，以及它们的成熟和糖基化。用p150,95特异性单克隆抗体（MAb）或针对变性、纯化的αX亚基的抗血清免疫沉淀p150,95α亚基。通过用亚基特异性MAb和抗血清进行免疫沉淀，以及用内切糖苷酶H和N-聚糖酶消化，比较了p150,95、Mac-1和淋巴细胞功能相关抗原（LFA-1）的α和β亚基的糖基化和多肽链长度。p150,95α亚基作为146,000 Mr的前体合成，有五到六个N-连接寡糖，多肽链主链为132,000 Mr。150,000 Mr成熟α亚基上超过50%的碳水化合物对内切糖苷酶H消化敏感。p150,95α和β前体在成熟为成熟形式之前可以结合。向成熟形式的转变伴随着与针对变性αX亚基的抗血清反应性的丧失，表明构象发生了变化。Mac-1和LFA-1α亚基的前体分别为160,000 Mr和165,000 Mr，并含有N-连接碳水化合物。Mac-1α亚基的多肽链长度为137,000 Mr，LFA-1的为149,000 Mr。成熟LFA-1α亚基上只有14%的寡糖对内切糖苷酶H敏感，这表明与p150,95不同，大多数寡糖转变为复合型。因此，三种同源α亚基在Mr上的差异是由于生物合成过程中多肽链长度和碳水化合物加工的差异。
