By contrast, in Rab5 the switch II aspartate is required for Rabex mediated GDP-release.
For DENND1-Rab35 anDrrA-Rab1b1 the Rab active-site glutamine, often mutated to create constitutively active forms, is involved in GEF mediated GDP-release.
The Rab RabGEF complex structures suggest this is possibly due to the negatively charged switch II aspartate, however mutation of this residue does not reduce GEF mediated GDP-release.
To better understand GEF-mediated GDP dissociation, we used computational approaches to recreate a putative on-pathway Arf1-GDP-GEF species immediately preceding nucleotide ejection, indicated in the schema above as intermediate II.
Circled residues are required for GEF-mediated GDP release.
The rate constant and the span of the intrinsic and the Cdc25-mediated GDP dissociation from C51S/C118S Ras were similar to that of the intrinsic and the Cdc25-mediated GDP dissociation from wt Ras.
This is equivalent to the minimal velocity (app Vmin) of the GEF-mediated GDP dissociation from small GTPases.
Intriguingly, although C118S Ras lacks Cys and wt Ras-SNO possesses the chemically modified redox inert Cys side chain, O2– was able to perturb the Cdc25-mediated GDP dissociation from C118S Ras and wt Ras-SNO.
Moreover, the values of the rate constant and span of the Cdc25-mediated GDP dissociation from C118S Ras and wt Ras-SNO were similar to those of the rate constant and span of the Cdc25-mediated wt Ras GDP dissociation.
Nevertheless, the values of the rate constants and the spans of the intrinsic, the Cdc25-mediated, and the O2 -mediated C51Ras GDP dissO2 -mediatede, respeC51Rasy, similar to those of the values of the intrinsic, Cdc25-mediated, and O2–-mediated wt Ras GDP dissociation.
When the oxidant concentration is fixed, not only the rate constant but also the span of the oxidant-mediated GTPase GDP dissociation may differ from the rate constant and the span of the GEF-mediated GTPase GDP dissociation.
