Armstrong S A, Hannah V C, Goldstein J L, Brown M S
Department of Molecular Genetics, University of Texas Southwestern Medical Center at Dallas 75235-9046, USA.
J Biol Chem. 1995 Apr 7;270(14):7864-8. doi: 10.1074/jbc.270.14.7864.
RhoB, a small GTP-binding protein, was shown previously to contain farnesyl (C-15) as well as geranylgeranyl (C-20) groups (Adamson, P., Marshall, C. J., Hall, A., and Tilbrook, P. A. (1992) J. Biol. Chem. 267, 20033-20038). The COOH-terminal sequence of the protein is CCKVL. According to current rules of prenylation, the COOH-terminal leucine should render the protein a substrate for CAAX geranylgeranyl transferase (GGTase-1), but not for CAAX farnesyltransferase (FTase). To determine the mechanism of farnesylation, we prepared recombinant RhoB and incubated it with recombinant preparations of either FTase or GGTase-1. RhoB was neither farnesylated nor geranylgeranylated efficiently by FTase, but it was farnesylated as well as geranylgeranylated by GGTase-1. The enzyme attached farnesyl more efficiently than geranylgeranyl to RhoB. Neither farnesylation nor geranylgeranylation required the cysteine at the fifth position from the COOH terminus. However, replacement of the cysteine at the fourth position abolished attachment of both prenyl groups. We conclude that the previously observed farnesylation of RhoB is attributable to the FTase activity of GGTase-1. These data, and other accumulating data, indicate that GGTase-1 is a highly unusual enzyme that efficiently transfers both farnesyl and geranylgeranyl groups and that the choice of prenyl group is dictated by the nature of the protein acceptor.
RhoB是一种小GTP结合蛋白，先前已证明其含有法尼基（C-15）以及香叶基香叶基（C-20）基团（亚当森，P.，马歇尔，C.J.，霍尔，A.，和蒂尔布鲁克，P.A.（1992）《生物化学杂志》267，20033 - 20038）。该蛋白的COOH末端序列为CCKVL。根据目前的异戊二烯化规则，COOH末端的亮氨酸应使该蛋白成为CAAX香叶基香叶基转移酶（GGTase - 1）的底物，但不是CAAX法尼基转移酶（FTase）的底物。为了确定法尼基化的机制，我们制备了重组RhoB，并将其与FTase或GGTase - 1的重组制剂一起孵育。FTase不能有效地将RhoB法尼基化或香叶基香叶基化，但GGTase - 1能将其法尼基化以及香叶基香叶基化。该酶将法尼基连接到RhoB上的效率高于香叶基香叶基。法尼基化和香叶基香叶基化都不需要COOH末端第五位的半胱氨酸。然而，替换第四位的半胱氨酸会消除两个异戊二烯基团的连接。我们得出结论，先前观察到的RhoB的法尼基化归因于GGTase - 1的FTase活性。这些数据以及其他积累的数据表明，GGTase - 1是一种非常特殊的酶，它能有效地转移法尼基和香叶基香叶基基团，并且异戊二烯基团的选择由蛋白受体的性质决定。
