Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/92215
標題: Stenotrophomonas maltophilia FleQ 及 Vibrio cholerae FlrA 蛋白的結構與功能分析
Structure and function studies of Stenotrophomonas maltophilia FleQ and Vibrio cholerae FlrA
作者: 施旻劭
Min-Shao Shih
關鍵字: 二級訊息傳遞子
cyclic di-GMP
FleQ
FleN
FlrA
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摘要: 過去的文獻指出FleQ 蛋白是控制細菌鞭毛基因表現的主要調控蛋白之一。它為一俱複雜功能的蛋白,可以做為一個 enhancer binding protein,也可以是一個 repressor protein。FleQ 蛋白的 N 端為 FleQ domain,中間為 AAA+/ATPase σ54 interaction domain,C 端為 helix-turn-helix DNA binding domain。另外,在文獻中也提到 FleQ 蛋白會受到 FleN 蛋白的調控而控制鞭毛的數量。FleN 蛋白為一個 ATPase,帶有一個特殊的 Walker A motif。在二級訊息傳遞分子 c-di-GMP 存在的情況下,FleN 蛋白上的 Walker A motif 會結合到 FleQ 蛋白的 N 端 domain (FleQ domain),進而調控 pel、psl 和 cdr 的基因表現,影響細菌合成鞭毛的蛋白表現,促使細菌產生生物膜 (biofilm)。 為了瞭解 FleQ 蛋白的作用方式,我們擬解析 FleQ 蛋白和與其功能相似的 FlrA 蛋白的結構。目前我們已經成功大量表現 S. maltophilia FleQ 全長蛋白、 N 端 FleQ domain 以及 Vibrio cholerae FlrA 全長和部分片段的蛋白。經由 X-光晶體繞射方法分析 SeMet-labeled SmFleQ domain 的蛋白晶體,解析度 (resolution) 可達 2.58 A,並且已成功解析出 SmFleQ domain 的結構。根據我們解析出的結構發現, SmFleQ domain 雖然被歸類為 REC domain,但是 SmFleQ domain 未帶有能接受磷酸根的 Asp,而且 SmFleQ domain 是以 α1-α5 相互作用形成二聚體,這些特點皆與典型的 REC domain 不同。本篇研究的結果對於非典型的 REC domain 有初步的了解。
FleQ is a unique enhancer binding protein regulating many genes involved in the bacterial flagellar and exopolysaccharide biosynthesis. By responding to c-di-GMP, it helps modulating the transition between planktonic and biofilm lifestyles in some well studied bacteria. FleQ is a multi-domain activator belonging to the NifA/NtrC enhancer binding protein (EBP) family and comprises an N-terminal FleQ domain, an AAA+/ATPase α54 interaction domain, and a helix-turn-helix DNA binding domain. Unlike other bacterial two component signal transduction systems that transmit signals via the phosphorelay of the REC domain, FleQ is regulated through the binding of an anti-activator protein FleN to its N-terminal FleQ domain. In this manuscript, we reported the 2.58 A resolution crystal structure of the Stenotrophomonas maltophilia (S. maltophilia) FleQ domain. It is found to belong to the REC domain family, but adopts many atypical feature, including the absence of the crucial Asp residue in the acidic binding pocket to accept a phosphoryl group from a upstream cognate histidine kinase,?dislocated α2/α3/α4 helices, and a unique α1-α5 dimeric interface. These results form a basis to further understand how noncanonical REC domain transmit environmental signals in the bacterial kingdom.
URI: http://hdl.handle.net/11455/92215
文章公開時間: 2017-10-20
Appears in Collections:生物化學研究所

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