Kaushansky K, O'Hara P J, Hart C E, Forstrom J W, Hagen F S
Division of Hematology, University of Washington, Seattle 98195.
Biochemistry. 1987 Jul 28;26(15):4861-7. doi: 10.1021/bi00389a038.
cDNA clones for the human hematopoietic regulator granulocyte-macrophage colony-stimulating factor (hGM-CSF) were isolated from a lamba gt11 cDNA library prepared from RNA of COS cells transiently expressing the gene for hGM-CSF. As the RNA was a rich source of hGM-CSF mRNA, approximately 0.1% of the clones of this library contained hGM-CSF sequences. All of the clones analyzed were full length and were correctly processed. When subcloned into an expression vector and transfected into COS cells, the cDNA clones direct the synthesis of higher levels of the growth factor than the gene from which they were derived. The cDNA for native hGM-CSF was used to generate structural mutants which lack N-linked carbohydrate, O-linked carbohydrate, or both. Although the mutant proteins had differing specific activities, the nonglycosylated forms reproduce many, if not all, of the physiologic functions of authentic hGM-CSF. The role of carbohydrate in the secretion and function of hGM-CSF is discussed.
从一个λgt11 cDNA文库中分离出了人造血调节因子粒细胞-巨噬细胞集落刺激因子（hGM-CSF）的cDNA克隆，该文库是用瞬时表达hGM-CSF基因的COS细胞的RNA构建的。由于该RNA是hGM-CSF mRNA的丰富来源，此文库中约0.1%的克隆含有hGM-CSF序列。所有分析的克隆均为全长且加工正确。当亚克隆到表达载体并转染到COS细胞中时，这些cDNA克隆指导合成的生长因子水平高于其来源基因。天然hGM-CSF的cDNA被用于产生缺乏N-连接碳水化合物、O-连接碳水化合物或两者皆缺的结构突变体。尽管突变蛋白具有不同的比活性，但非糖基化形式重现了天然hGM-CSF许多（即便不是全部）生理功能。文中讨论了碳水化合物在hGM-CSF分泌和功能中的作用。
