Trapped!

Small G proteins (for example, Arfs) are activated when guanine nucleotide exchange factors (GEFs; for example, ARNO) catalyse the exchange of GDP for GTP. This process involves G-protein–GDP–GEF intermediates, but, owing to their transient nature, the structures of these intermediates have remained a mystery. Now, though, Cherfils and colleagues provide new insights in Nature.

Arf proteins regulate membrane trafficking in eukaryotic cells, and they couple their GDP/GTP exchange to cytosol–membrane translocation. Transient GDP-bound intermediates of these G proteins can be trapped either by the natural inhibitor brefeldin A, or by a mutation in the catalytic Sec7 domain of their GEFs (E156K in ARNO). The authors therefore determined the crystal structures of these intermediates, and found that they seem to represent two actual intermediates of the exchange reaction — that is, a G-protein–GDP–GEF docking intermediate and a pre-GDP-dissociation intermediate. The structures indicate that the Sec7 domain catalyses the membrane recruitment of Arf and the dissociation of GDP from Arf as separate, sequential reactions, and that these events involve the sequential rotation of the Arf–GDP core towards the Sec7 catalytic site. The two structures represent unproductive G-protein–GEF complexes and have highlighted “...how efficient and selective inhibition of a cellular process in vivo can be achieved by a drug that targets a transient protein–protein intermediate of a regulatory reaction and traps it in a non-productive conformation”. REFERENCE Renault, L. et al. Structural snapshots of the mechanism and inhibition of a guanine nucleotide exchange factor. Nature 426, 525–530 (2003)

A refined model

The Sec61 (eukaryotes) or SecY (eubacteria and archaea) complex is a protein-conducting channel that allows polypeptides to cross, or integrate into, membranes. It's a heterotrimeric complex with α-, β- and γ-subunits, and the α-subunit forms the pore of the channel. The channel is passive and it associates with other components that provide the driving force for translocation. Only relatively low-resolution structures of this complex have been available to date, but now, in Nature, Rapoport and colleagues describe the 3.2-Å resolution crystal structure of the SecY complex from Methanococcus jannaschii. This structure, combined with published data, has allowed the authors to propose models for the various steps of protein translocation.

The structure indicates that a single copy of SecY functions as a channel, despite the fact that this complex forms oligomers during translocation. A cytoplasmic funnel that leads into the channel is plugged by a short helix, and the authors propose that displacement of this plug opens the channel. The channel is hourglass shaped with a 'pore ring' of hydrophobic residues at its narrowest point. These residues might form a seal around the translocating polypeptide to block the passage of other molecules. The structure also indicates how the complex interacts with the polypeptide's signal sequence and the components that provide the driving force. Furthermore, it shows how the transmembrane segments of nascent membrane proteins might move laterally into lipid, and has provided the basis for future experimental testing. REFERENCE van den Berg, B. et al. X-ray structure of a protein-conducting channel. Nature 3 Dec 2003 (doi:10.1038/nature02218)