Acta Crystallographica Section D

Biological Crystallography

Volume 69, Part 3 (March 2013)

research papers

Acta Cryst. (2013). D69, 345-351    [ doi:10.1107/S0907444912047294 ]

Direct metal recognition by guanine nucleotide-exchange factor in the initial step of the exchange reaction

T. Uejima, K. Ihara, M. Sunada, M. Kawasaki, T. Ueda, R. Kato, A. Nakano and S. Wakatsuki

Abstract: Rab small GTPases regulate vesicle transport in eukaryotes by interacting with various effectors. Guanine nucleotide-exchange factor (GEF) catalyzes the transition from inactive GDP-bound Rab to active GTP-bound Rab. The existence of several GDP-bound intermediates containing the Arabidopsis thaliana Rab5 homologue ARA7 and the GEF VPS9a prior to the formation of a nucleotide-free binary complex has been proposed [Uejima et al. (2010), J. Biol. Chem. 285, 36689-36697]. During this process, VPS9a directly interacts with the [beta]-phosphate of GDP and the P-loop lysine of ARA7 via a catalytically important aspartate finger, which promotes the release of GDP from ARA7. However, it is unclear how VPS9a removes Mg2+ from ARA7 before forming the GDP-bound ternary complex. Here, the structure of the ARA7-GDP-Ca2+-VPS9a complex is reported, in which the aspartate finger directly coordinates the divalent metal ion. Ca2+ is bound to the canonical Mg2+-binding site, coordinated by the [beta]-­phosphate of GDP and the P-loop serine of ARA7. Unexpectedly, Ca2+ is further coordinated by the aspartate finger and the main chain of VPS9a. This structure may represent the earliest intermediate step in the GEF-catalyzed nucleotide-exchange reaction of ARA7 before the metal-free GDP-bound intermediates are created.

PDB reference: 4g01

Keywords: guanine nucleotide-exchange factors; ARA7.

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