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Crystal structure of the Sec4p·Sec2p complex in the nucleotide exchanging intermediate state

Sato, Yusuke, Fukai, Shuya, Ishitani, Ryuichiro, Nureki, Osamu
Proceedings of the National Academy of Sciences of the United States of America 2007 v.104 no.20 pp. 8305-8310
crystal structure, exocytosis, guanine nucleotides, guanosinetriphosphatase, hydrophobicity, phosphates, regulatory proteins, yeasts
Vesicular transport during exocytosis is regulated by Rab GTPase (Sec4p in yeast), which is activated by a guanine nucleotide exchange factor (GEF) called Sec2p. Here, we report the crystal structure of the Sec2p GEF domain in a complex with the nucleotide-free Sec4p at 2.7 Å resolution. Upon complex formation, the Sec2p helices approach each other, flipping the side chain of Phe-109 toward Leu-104 and Leu-108 of Sec2p. These three residues provide a hydrophobic platform to attract the side chains of Phe-49, Ile-53, and Ile-55 in the switch I region as well as Phe-57 and Trp-74 in the interswitch region of Sec4p. Consequently, the switch I and II regions are largely deformed, to create a flat hydrophobic interface that snugly fits the surface of the Sec2p coiled coil. These drastic conformational changes disrupt the interactions between switch I and the bound guanine nucleotide, which facilitates the GDP release. Unlike the recently reported 3.3 Å structure of the Sec4p·Sec2p complex, our structure contains a phosphate ion bound to the P-loop, which may represent an intermediate state of the nucleotide exchange reaction.