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Structural Analysis of Pin1 and Its Homologues

Author: SunLiFang
Tutor: LiuYiCheng；WuXueJi
School: Xiamen University
Course: Biochemistry and Molecular Biology
Keywords: Pin1 Circular dichroism Crystallization Structure
CLC: Q51
Type: Master's thesis
Year: 2009
Downloads: 83
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Abstract

The protein proline in front of the serine or threonine reversible phosphorylation is an important regulation of cell signaling mechanisms . Ser / Thr - proline motif recently discovered peptide prolyl cis-trans isomerase Pin1 can specifically recognize and bind to protein phosphorylation , catalyzed phosphorylation of Ser / Thr - proline peptide bond cis-trans isomers , thereby changing the phosphorylation function of the protein . More and more research data indicate that this configuration change affects the catalytic activity of the protein , the interaction between the level of protein and protein dephosphorylation , the subcellular localization of the protein and protein transporter . Moreover, the adjustment mechanism of this phosphorylation in cell growth and disease occurs to play an important role , such as cancer and Alzheimer's disease . Pin1 catalytic adjustment mechanism , as well as its interaction with the substrate . This thesis on two key domains of the human Pin1 protein Pin1 several key amino acids Trp34 , Lys63 , Cys113 , Met130 directed mutagenesis , we a lot of expression of these proteins , separation , purification , and then suspended liquid drop vapor diffusion prepared crystals by rocking method to collect diffraction data and crystal structure analysis . We get the Human Pin1-WT, Pin1-W34A, Pin1-K63A, Pin1-C113A, Pin1-M130A. The research results show that the mutant Pin1-W34A phosphorylation of the substrate specificity of the ability to identify and catalytic function in vitro loss of Pin1 - K63A mutant Pin1 - C113A substrate phosphorylation - specific recognition unaffected However , the loss of its catalytic function ; same time, we found from the Pin1-K63A, Pin1-C113A and Pin1-M130A PPIase domain there may be the root of the two phosphate binding sites . We successfully cloned , expressed Arabidopsis Pin1 ( AtPin1 ) , contains only the PPIase domain of E.coli Pin1 (E.coli Par10) and Trypanosoma brucei Pin1 ( TbPin1 ) , and has been AtPin1 E.coli Par10 crystal , but AtPinl and E.coli Par10 crystal quality is not high , need to optimize ; while TbPin1 the not yet obtained crystal . The study was to lay the foundation for the further study of the mechanism of Pin1 and structure theoretical basis for future Pin1 - based drug design .

Structural Analysis of Pin1 and Its Homologues

Abstract

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