Crystallization and crystallographic analysis of yeast Sec2p, a guanine nucleotide-exchange factor for the yeast Rab GTPase Sec4p

Sec2p is a guanine nucleotide-exchange factor (GEF) for the yeast Rab GTPase Sec4p. Sec2p accelerates GDP release from Sec4p and promotes GDP-GTP exchange for Sec4p activation. In order to elucidate this nucleotide-exchange mechanism using X-ray crystallography, three constructs of native Sec2p (Sec2_1-160p, Sec2_18-160p and Sec2_31-160p) and three constructs of selenomethionine-labelled Sec2p [Sec2_31-160p, Sec2_31-160p (M115L) and Sec2_31-160p (M115L, K121M, T142M)] were crystallized. These six crystals diffracted to 8.8, 4.8, 2.6, 4.0, 3.3 and 3.0 Å resolution, respectively. The data set from the SeMet-labelled Sec2_31-160p (M115L, K121M, T142M) crystal was processed for SAD phasing; the crystal belonged to space group P2₁2₁2₁, with unit-cell parameters a = 101.9, b = 176.6, c = 181.5 Å.