A Tethered Catalysis, Two - Hybrid System to Identify Protein-protein Interactions Requiring Post Translational Modifications  

Oleh:

Dawei Guo ; Hazbun, Tony R. ; Xin-Jing, Xu ; Sze-Ling, Ng ; Fields, Stanley ; Min-Hao Kuo

Jenis: Article from Journal - ilmiah internasional
Dalam koleksi: Nature Biotechnology: The Science and Business of Biotechnology vol. 22 no. 7 (Jul. 2004), page 888-892.
Topik: PROTEIN; tethered catalysis; two hybrid system; protein interaction; post translational modification

Ketersediaan

Perpustakaan Pusat (Semanggi) Nomor Panggil: NN9.1 Non-tandon: 1 (dapat dipinjam: 0) Tandon: tidak ada

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Isi artikel

We have modified the yeast two hybrid system to enable the detection of protein-protein interactions that require a specific post translational modification, using the acetylation of histones and the phosphorylation of the carbocyl terminal domain (CTD) of RNA polymerase II as test modifications. In this testhered catalysis assay, constitutive modification of the protein to be sreneed for interactions is achieved by fusing it to its cognate modifying enzyme, with the physical linkage resulting in eggicient catalysis. This catalysis maintain substrate modification even in the presence of antagonizing enzyme activities. A catalitically inactive mutant of the enzyme is fused to the substrate as a control such that the modification does not occur, this construct enables the rapid identification of modification independent interactions. We identified proteins with links to chromatin functions that interact with acetylated histones, and proteins that participate in RNA polymerase II functions and in CTD phosphorylation regulation that interact preferentially with the phosphorylated CTD.

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