Figure 2.

Geranylgeranylation of recombinant Myc-Rab1B. (A) Aliquots of E. coli lysate containing 20–30 pmol of each of the indicated recombinant Myc-tagged Rab1B proteins were geranylgeranylated in cell-free reactions as described in MATERIALS AND METHODS. The top portion shows an autoradiograph of the ¹²⁵I-labeled immunoblot (18-h exposure) containing half of each prenylation reaction, confirming that comparable amounts of Myc-Rab1B were present in the assays. The bottom portion shows the corresponding fluorograph (24-h exposure) of the other half of the reaction used for determination of [³H]GG incorporation. The ³H:¹²⁵I ratios determined for each Rab1B construct are summarized in Table 1. (B) Aliquots of E. coli lysate containing approximately 10 pmol of recombinant Rab1B expressed with or without the amino-terminal Myc epitope tag were geranylgeranylated in parallel reactions and the proteins were subjected to SDS-PAGE and fluorography (72-h exposure).