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. Author manuscript; available in PMC: 2010 Apr 1.
Published in final edited form as: Mol Microbiol. 2009 Mar 6;72(2):442–458. doi: 10.1111/j.1365-2958.2009.06657.x

Fig. 2.

Fig. 2

In vitro kinase assay of TbAUK1.

(A) Constitutive expression of AU1-tagged TbAUK1. Parental AnTat1.1 PF were transformed with the constitutive expression vector pTSA-AU1.TbAUK. When pulled down with anti-AU1 Sepharose beads, transformants phosphorylated myelin basic protein (MBP), in a manner that was inhibited by Hesperadin. Parental cells phosphorylated MBP at a background level.

(B) Inducible expression of AU1-tagged wild-type TbAUK1 and the K58R mutated TbAUK1. The tagged proteins were detected in cell homogenates by western blot with antibodies against AU1 (upper panels). TbRACK1 was used as a loading control (middle panels). The immunoprecipitated proteins phosphorylated MBP (lower panel).

(C) Nucleotide specificity of TbAUK1. Unlabeled nucleotides (1 mM each) were added to the standard reaction mix. Only unlabeled ATP prevented phosphorylation of MBP.