Brenner M, Lampel K, Nakatani Y, Mill J, Banner C, Mearow K, Dohadwala M, Lipsky R, Freese E
National Institutes of Health, National Institute of Neurological Disorders and Stroke, Laboratory of Molecular Biology, Bethesda, MD 20892.
Brain Res Mol Brain Res. 1990 May;7(4):277-86. doi: 10.1016/0169-328x(90)90078-r.
Both a partial cDNA clone and a complete genomic clone have been isolated for human gfa, the gene encoding the major component of astrocyte intermediate filaments, glial fibrillary acidic protein (GFAP). The nucleotide sequence of the entire coding region and 102 bp of the 5' flanking DNA was determined. The mRNA start site was identified by primer extension and probe protection experiments, and a novel in vitro transcription and translation procedure was then used to establish that the first ATG in the mRNA initiates GFAP synthesis. The predicted amino-terminal sequence for human GFAP differs greatly from that previously deduced for mouse GFAP from its gene sequence, despite otherwise high homology. This discrepancy was resolved by determining that the published mouse genomic sequence has an incorrect additional base. The corrected sequence produces strong homology between human and mouse GFAP in their predicted amino acid sequences, and suggests that human and mouse GFAP initiate at homologous positions. The beginning sequence deduced here for both proteins is matched closely by that previously obtained for porcine GFAP by direct sequencing of its amino-terminal end. This supports the protein initiation sites proposed, and also indicates that GFAP is not processed at its amino-terminal end.
人类胶质纤维酸性蛋白（GFAP）的部分cDNA克隆和完整基因组克隆均已分离出来，GFAP是编码星形胶质细胞中间丝主要成分的基因。测定了整个编码区及5'侧翼DNA的102 bp的核苷酸序列。通过引物延伸和探针保护实验确定了mRNA起始位点，然后采用一种新的体外转录和翻译方法来确定mRNA中的第一个ATG启动GFAP的合成。尽管人类GFAP与小鼠GFAP在其他方面具有高度同源性，但其预测的氨基末端序列与先前根据小鼠GFAP基因序列推导的序列有很大差异。通过确定已发表的小鼠基因组序列有一个错误的额外碱基，解决了这一差异。校正后的序列在人类和小鼠GFAP的预测氨基酸序列之间产生了很强的同源性，并表明人类和小鼠GFAP在同源位置起始。这里推导的两种蛋白质的起始序列与先前通过直接测序猪GFAP氨基末端获得的序列非常匹配。这支持了所提出的蛋白质起始位点，也表明GFAP在其氨基末端不进行加工。
