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Biochemical and structural characterization of MUPP1-PDZ4 domain from Mus musculus

Biochemical and structural characterization of MUPP1-PDZ4 domain from Mus musculus
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摘要 特定的 proteinprotein 相互作用为生物信号 transduction 是重要的。postsynaptic density-95,圆盘大,并且 zonulin-1 (PDZ ) 领域是最丰富的蛋白质相互作用模块之一。(MUPP1 ) 作为支架蛋白质, Multi-PDZ-domain 蛋白质 1 包含 13 个 PDZ 领域并且在细胞骨架组织,房间极性,和房间增长起一个重要作用。MUPP1 的 PDZ 领域上的学习帮助理解 MUPP1 的机制和功能。在现在的学习,从亩 musculus 的 MUPP1 (MUPP1-PDZ4 ) 的第四个 PDZ 领域被克隆,表示,净化,并且描绘。MUPP1-PDZ4 领域是进宠物向量的 subcloned 并且在 Escherichia 表示了 coli。亲密关系层析和尺寸排除层析被用来净化蛋白质。MUPP1-PDZ4 蛋白质是有在答案的 16.4 kDa 的一个臼齿的团的单体并且让融化 60.3he 削尖 diffe 褄?褄 的主要决定因素吗?? Specific protein-protein interactions are important for biological signal transduction. The postsy- naptic density-95, disc-large, and zonulin-1 (PDZ) domain is one of the most abundant protein inter- action modules. Multi-PDZ-domain protein 1 (MUPP1), as a scaffold protein, contains 13 PDZ domains and plays an important role in cytoskeletal organization, cell polarity, and cell prolifer- ation. The study on PDZ domain of MUPP1 helps to understand the mechanisms and functions of MUPPI. In the present study, the fourth PDZ domain of MUPP1 (MUPP1-PDZ4) from Mus musculus was cloned, expressed, purified, and characterized. The MUPP1-PDZ4 domain was sub- cloned into a pET-vector and expressed in Escherichia coil Affinity chromatography and size- exclusion chromatography were used to purify the protein. MUPP1-PDZ4 protein was a monomer with a molar mass of 16.4 kDa in solution and had a melting point of 60.3~C. Using the sitting-drop vapor-diffusion method, MUPP1-PDZ4 protein crystals were obtained in a solution (pH 7.0) contain- ing 2% (v/v) polyethylene glycol 400, 0.1 M imidazole, and 24% (w/v) polyethylene glycol mono- ethyl ether 5000. Finally, the crystal was diffracted with 1.6A. resolution. The crystal structure showed that MUPP1-PDZ4 domain contained three m-helices and six 13-strands in the core. The GLGI motif, L562/A564 on the B-strand B, and H605N608/L612 on the a-helix B formed a PDZ bind- ing pocket which could bind to the C-terminal of the binding partners. This biochemical and struc- tural information will provide insights into how PDZ binds to its target peptide and the theoretical foundation for the function of MUPPI.
作者 Haili.Zhu Zexu Liu Yuxin Huang ChaoZhang Gang Li Wei Liu Haili Zhu1 ;Zexu Liu2 ;Yuxin Huang1 ;Chao Zhang1 ;Gang Li3; Wei Liu1,2,*
出处 《生物化学与生物物理学报:英文版》 SCIE CAS CSCD 2015年第3期199-206,共8页 Acta Biochimica et Biophysica Sinica
基金 We thank the BL17U1 beamline team of the Shanghai Synchrotron Radiation Facility for the X-ray beam time.
作者简介 Correspondence address. Tel/Fax: +86-755-83910721; E-mail: liuw@ust.hk
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