Asymmetric Usage of Antagonist Charged Residues Drives Interleukin-5 Receptor Recruitment but Is Insufficient for Receptor Activation

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• 作者：Tetsuya Ishino ; Udaya Pillalamarri ; Dominick Panarello ; Madhushree Bhattacharya ; Cecilia Urbina ; Stephanie Horvat ; Sanjay Sarkhel ; Bradford Jameson ; and Irwin Chaiken
• 刊名：Biochemistry
• 出版年：2006
• 出版时间：January 31, 2006
• 年：2006
• 卷：45
• 期：4
• 页码：1106 - 1115
• 全文大小：327K
• 年卷期：v.45,no.4(January 31, 2006)
• ISSN：1520-4995

文摘

The cyclic peptide AF17121 (VDECWRIIASHTWFCAEE) is a library-derived antagonist forhuman Interleukin-5 receptor mages/gifchars/alpha.gif" BORDER=0> (IL5Rmages/gifchars/alpha.gif" BORDER=0>). We have previously demonstrated that AF17121 mimicsInterleukin-5 (IL5) by binding in a region of IL5Rmages/gifchars/alpha.gif" BORDER=0> that overlaps the IL5 binding epitope. In the presentstudy, to explore the functional importance of the amino acid residues of AF17121 required for effectivebinding to, and antagonism of, IL5Rmages/gifchars/alpha.gif" BORDER=0>, each charged residue was subjected to site-directed mutagenesisand examined for IL5Rmages/gifchars/alpha.gif" BORDER=0> interaction by using a surface plasmon resonance biosensor. One residue, Arg⁶,was found to be essential for receptor antagonism; its replacement with either alanine or lysine completelyabolished the interaction between AF17121 and IL5Rmages/gifchars/alpha.gif" BORDER=0>. Other charged residues play modulatory roles.One class consists of the N-terminal acidic cluster (Asp² and Glu³) for which alanine replacement decreasedthe association rate. A second class consists of His¹¹ and the C-terminal acidic cluster (Glu¹⁷ and Glu¹⁸)for which alanine replacement increased the dissociation rate. Binding model analysis of the mutants ofthe latter class of residues indicated the existence of conformational rearrangement during the interaction.On the basis of these results, we propose a model in which Arg⁶ and N-terminal acidic residues drive theencounter complex, while Arg⁶, His¹¹, and C-terminal acidic residues are involved in stabilizing the finalcomplex. These data argue that the charged residues of AF17121 are utilized asymmetrically in the pathwayof inhibitor-receptor complex formation to deactivate the receptor function. The results also help focusemerging models for the mechanism by which IL5 activates the IL5Rmages/gifchars/alpha.gif" BORDER=0>-mages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">c receptor system.