Purification, crystallization and initial X-ray crystallographic analysis of the putative GTPase PH0525 from Pyrococcus horikoshii OT3
Neratur Krishnappagowda Lokanath,Hitoshi Yamamoto,Emiko Matsunaga,Mitsuaki Sugahara,Naoki Kunishima +4 more
TL;DR: The putative GTPase PH0525 from Pyrococcus horikoshii OT3 has been overexpressed in Escherichia coli and purified and is consistent with dynamic light-scattering experiments, which show a monomeric state of the protein in solution.
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Abstract: The putative GTPase PH0525 from P. horikoshii OT3 was crystallized using the microbatch method. Crystals were formed under two different conditions, providing two distinct crystal forms. Diffraction data from the two forms were measured to resolution limits of 2.30 and 2.40 A and processed in space groups P2{sub 1} and C222{sub 1}, respectively. GTPases are involved in diverse cellular functions including cell proliferation, cytoskeleton organization and intracellular traffic. The putative GTPase PH0525 from Pyrococcus horikoshii OT3 has been overexpressed in Escherichia coli and purified. Two distinct crystal forms were grown by the microbatch method at 291 K using a very high protein concentration (80 mg ml{sup −1}). Native data sets extending to resolutions of 2.3 and 2.4 A have been collected and processed in space groups P2{sub 1} and C222{sub 1}, respectively. Assuming the presence of one monomer per asymmetric unit gives V{sub M} values of 2.6 and 2.4 A{sup 3} Da{sup −1} for the P2{sub 1} and C222{sub 1} forms, respectively, which is consistent with dynamic light-scattering experiments, which show a monomeric state of the protein in solution.
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