Crystallization and preliminary X-ray studies of mouse centrin1

Acta Crystallographica. Section F, Structural Biology and Crystallization Communications
Jung Hee ParkHui-Woog Choe

Abstract

Centrins belong to a family of Ca2+-binding EF-hand proteins that play a fundamental role in centrosome duplication and the function of cilia. To shed light on the structure-function relationship of these proteins, mouse centrin1 has been crystallized. The mouse centrin1 has been expressed in Escherichia coli as a GST-centrin fusion protein containing a thrombin protease cleavage site between the fusion partners. Two constructs with different linking-sequence lengths were expressed and purified. Thrombin cleavage yielded functional centrin1 and N-terminally extended centrin1 containing 25 additional residues upstream of its N-terminus. Only N-terminally extended centrin1 (MW approximately 22 240 Da) could be crystallized at room temperature, using 20-25%(w/v) PEG 1500, 5-10%(v/v) ethylene glycol and 1-2%(v/v) dioxane. Crystals were suitable for X-ray analysis, diffracting to 2.9 A at 295 K using a rotating-anode X-ray source. They belong to space group C2, with unit-cell parameters a = 60.7, b = 59.6, c = 58.3 A, beta = 109.4 degrees. Assuming the asymmetric cell to be occupied by one centrin1 molecule of 22.2 kDa, the unit cell contains 45% solvent with a crystal volume per protein weight, VM, of 2.2 A3 Da(-1).

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Citations

Jul 6, 2006·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Jean Baptiste CharbonnierConstantin T Craescu
Mar 11, 2008·Progress in Retinal and Eye Research·Philipp TrojanUwe Wolfrum
May 29, 2016·Protein Expression and Purification·Regur PhanindranathYogendra Sharma
Oct 19, 2017·Biochemistry. Biokhimii︠a︡·So Young KimJung Hee Park

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