Crystallization and preliminary crystallographic analysis of the circadian clock protein KaiB from the thermophilic cyanobacterium Thermosynechococcus elongatus BP-1

Abstract

KaiB is a component of the circadian clock oscillator in cyanobacteria, which are the simplest organisms that exhibit circadian rhythms. KaiB consists of 108 amino-acid residues and has a molecular weight of 12 025 Da. KaiB and Cys-substituted KaiB mutants from the thermophilic cyanobacterium Thermosynechococcus elongatus BP-1 were expressed as GST-fusion proteins in Escherichia coli, purified and crystallized. The crystals of wild-type KaiB belong to the monoclinic space group P21, with unit-cell parameters a = 89.6, b = 71.2, c = 106.8 Å, β = 100.1°. While the native crystals diffract to 3.7 Å, osmium derivatives, which show an approximately 4 Å shrinkage in the b axis, diffract to 2.6 Å. The crystals of the singly Cys-substituted mutant T64C with Hg, which show different morphology, diffract to 2.5 Å and belong to the monoclinic space group P2, with unit-cell parameters a = 63.7, b = 33.4, c = 93.7 Å, β = 100.1°. Anomalous difference Patterson maps of the Os- and Hg-derivative crystals had significant peaks in their Harker sections, suggesting that both derivatives are suitable for structure determination. © 2004 International Union of Crystallography. Printed in Denmark - all rights reserved.