The Interaction Between Tec Family Protein Tyrosine Kinase Itk PH Domain and OS-9 Protein

HAN Yue heng, ZHANG Wen hong, ZHANG Xiao guang, LIU Xin ping, YAO Li bo 

Chinese Journal of Biochemistry and Molecular Biology ›› 2003, Vol. 19 ›› Issue (04) : 499-503.

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Chinese Journal of Biochemistry and Molecular Biology ›› 2003, Vol. 19 ›› Issue (04) : 499-503.
Article

The Interaction Between Tec Family Protein Tyrosine Kinase Itk PH Domain and OS-9 Protein

  • HAN Yue heng, ZHANG Wen hong, ZHANG Xiao guang, LIU Xin ping, YAO Li bo 
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Abstract

The yeast two hybrid system was used to explore the partner of inducible T cell kinase (Itk) and the role of Itk in T cell signaling were investigated. Itk PH domain fragments were amplified by PCR using cloned Itk as template. The PH domain fragment was then cloned into a pLexA vector, which contained the DNA binding (BD) domain. The recombinant was used as a bait to screen a T cell cDNA library after it was testified to have neither ability of autonomous reporter gene activation, nor yeast cell toxicity. The genes of PH domain and the positive library cDNA fragment were cloned into prokaryotic expression vectors pGEX4T 1 and pET28a, respectively. And using the soluble fusion proteins an in vitro binding assay was employed to verify their interaction. One of the positive clones, sequenced and analyzed in the BLAST, was confirmed to be the gene fragment of OS 9, a protein which exists in many tissues while expressed highly in some sarcomas. The binding assay of the two fusion proteins demonstrated that they interacted with each other in vitro . OS 9 protein may be a potential partner of the Itk PH domain. The significance of the interaction between these two proteins is to be studied.

Key words

Itk / PH domain / yeast two hybrid system / fusion expression

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HAN Yue heng, ZHANG Wen hong, ZHANG Xiao guang, LIU Xin ping, YAO Li bo . The Interaction Between Tec Family Protein Tyrosine Kinase Itk PH Domain and OS-9 Protein[J]. Chinese Journal of Biochemistry and Molecular Biology, 2003, 19(04): 499-503

References

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