James G L, Goldstein J L, Pathak R K, Anderson R G, Brown M S
Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas 75235-9046.
J Biol Chem. 1994 May 13;269(19):14182-90.
CAAX farnesyltransferase attaches a farnesyl group to proteins that terminate in the sequence CAAX, where C is cysteine, A is an aliphatic amino acid, and X is typically methionine or serine. A limited number of substrates for the CAAX farnesyltransferase have been identified in cultured cells. These include p21ras proteins and the nuclear lamins A and B. We describe here the use of a CAAX farnesyltransferase inhibitor, together with a hamster cell line that exhibits efficient uptake of [3H]mevalonate, as a means of identifying novel farnesylated proteins. One candidate protein was purified and its attached prenyl group identified as farnesyl. The predicted amino acid sequence of this protein, deduced from a cloned cDNA, terminates with the tetrapeptide Cys-Leu-Ile-Met, which conforms to the consensus sequence for recognition by farnesyltransferase. This farnesylated protein, designated PxF, is localized to the outer surface of peroxisomes as determined by indirect immunofluorescence and electron microscopy.
CAAX法尼基转移酶将法尼基基团连接到以CAAX序列结尾的蛋白质上，其中C代表半胱氨酸，A代表脂肪族氨基酸，X通常为甲硫氨酸或丝氨酸。在培养细胞中已鉴定出有限数量的CAAX法尼基转移酶底物。这些底物包括p21ras蛋白以及核纤层蛋白A和B。我们在此描述了使用一种CAAX法尼基转移酶抑制剂，以及一种能高效摄取[3H]甲羟戊酸的仓鼠细胞系，作为鉴定新型法尼基化蛋白的一种方法。一种候选蛋白被纯化，其连接的异戊二烯基团被鉴定为法尼基。从克隆的cDNA推导得到的该蛋白的预测氨基酸序列以四肽Cys-Leu-Ile-Met结尾，这与法尼基转移酶识别的共有序列相符。通过间接免疫荧光和电子显微镜确定，这种法尼基化蛋白命名为PxF，定位于过氧化物酶体的外表面。
