Crystallization and preliminary X-ray diffraction study of the catalytic subunit of archaeal H⁺-transporting ATP synthase fromPyrococcus horikoshiiOT3

Abstract

H+ -transporting ATP synthase (H+ -ATPase) is a multi-subunit complex which acts to produce ATP molecules. The catalytic subunit A of the archaeal-type H+ -ATPase from Pyrococcus horikoshii OT3 was cloned, expressed in Escherichia coli, purified and crystallized by the hanging-drop vapour-diffusion method with MPD as a precipitant. X-ray intensity data were collected to 2.55 A resolution at beamline BL41XU of SPring-8. The crystals belong to the tetragonal space group P4(1)2(1)2 or P4(3)2(1)2, with unit-cell parameters a = b = 128.0, c = 104.7 A, and contain one molecule per asymmetric unit.

Nucleotide Binding States of Subunit A of the A-ATP Synthase and the Implication of P-Loop Switch in Evolution

“…22 Contaminating proteins were removed using the anion-exchange column Resource Q (6 ml), followed by gel filtration on the 26/60 Superdex200 column (GE Healthcare). The purity of the proteins was ascertained by SDS gel (Fig.…”

“…22 The proteins have been crystallized from a solution containing 50% (vol/vol) MPD and 0.1 M acetate (pH 4.5) at 18°C. Paraffin oil and silicone oil (vol/vol 1:1) were overlaid onto the mother liquor to a final volume of 200 μl.…”

Conservation of intron and intein insertion sites: implications for life histories of parasitic genetic elements

“…Positions of spliceosomal introns from are indicated as green dots without arrows. Panel B shows the structure of ATPase catalytic subunit A structure from Pyrococcus horikoshii OT3 (PDB ID: 1VDZ[68]) colored according to sequence conservation. The arrows indicate "a" and "b" intein insertion sites.…”

Crystal Structure of the Archaeal A1AO ATP Synthase Subunit B from Methanosarcina mazei Gö1: Implications of Nucleotide-binding Differences in the Major A1AO Subunits A and B

“…38 Since a Gln residue occupies a position equivalent to the Glu residue in the a subunits, it has been argued that the absence of the catalytic carboxylate in the corresponding loop would explain the lack of catalytic activity. 13 62 were fitted into a 23 Å electron density of the intact A-ATP synthase from T. thermophilus obtained by 3D reconstruction of negatively stained particles using the program Emfit. 25 The fitted coordinates of bovine mitochondrial F-ATP synthase g subunit (orange, PDB 1e1q, chain G).…”