Question

A researcher is conducting an experiment where they are monitoring vesicle assembly. Predict what effects on vesicle assembly they would see if the cells they are monitoring are exposed to non-hydrolyzable GTP.

Answer 1

            The assembly of vesicle coats also requires GTP-binding proteins, which appear to regulate the binding of coat proteins to the membrane. The budding of both clathrin-coated and COPI-coated vesicles from the Golgi complex requires a GTP-binding protein called ARF (ADP-ribosylation factor), while the budding of COPII-coated vesicles from the ER requires a distinct GTP-binding protein called Sar1. The role of these proteins is illustrated by the function of ARF in assembly of COPI-coated vesicles.

The first step in vesicle formation is the association of ARF bound to GDP with the Golgi membrane. Proteins in the Golgi membrane then stimulate the exchange of the GDP bound to ARF for GTP, and the COPI coat proteins bind to the ARF/GTP complex. Assembly of the coat is then followed by deformation of the membrane and vesicle budding. ARF then hydrolyzes its bound GTP, leading to the conversion of ARF to the GDP-bound state and the dissociation of coat proteins from the vesicle membrane.

            If GTP will not be hydrolysed then the vesicle will not be delivered to its target because the vesicle, coat protein and the ARF-GTP complex will not dissociate. No more ARF-GDP complexes will be available to initiate the process of coating the new vesicle.