Activity of the E75E76 mutant of the α subunit of casein kinase II from Xenopus laevis

FEBS Letters

Jedlicki, Ana¹  Allende, Catherine C.¹  Gatica, Marta¹  Allende, Jorge E.¹ 

[1]Departamento de Bioquímica, Facultad de Medicina, Universidad de Chile, Casilla 70086, Santiago 7, Chile

关键词: Casein kinase II;    Mutated α subunit;    Xenopus laevis;    Heparin;    Poly U;    Quercetin;   

DOI  :  10.1016/0014-5793(94)80392-7

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The cDNA gene coding for the α subunit of Xenopus laevis casein kinase II was mutated using the overlap extension PCR method. The mutation substituted glutamic acids for Lys⁷⁵ and Lys⁷⁶, changing the charge distribution of a very basic sequence found in the α subunit. Expression of the mutated cDNA in a pT7-7 vector in E. coli yielded an active mutant recombinant protein that was extensively purified. This mutant was not significantly affected in its app. K _m for casein or a model peptide substrate, nor in its interaction with the activating β subunit. Inhibition by quercetin and by 5,6-dichloro-1-β-d-ribofuranosyl benzimidazole was also the same for mutant and wild type subunits. However, the CKII αE⁷⁵E⁷⁶ mutant was at least one order of magnitude less sensitive to inhibition by polyanionic inhibitors such as heparin, poly U, copolyglutamic acid:tyrosine (4:1) and 2,3 diphosphoglycerate.

【 授权许可】

Unknown