生物谷報道：臺灣中研院生化所的王惠鈞（Andrew H.-J.Wang）和王廷方(Ting-Fang Wang)博士領(lǐng)導的一個研究組發(fā)現(xiàn)RecA家族重組酶功能是充當DNA損傷修復的一種新型的回轉(zhuǎn)馬達蛋白質(zhì),。研究組近期發(fā)表了兩篇有關(guān)RecA家族重組酶的結(jié)構(gòu)生物學相關(guān)文章。一篇文章發(fā)表9月12日的網(wǎng)絡(luò)版雜志PLoS ONE上,，另外一篇發(fā)表在今年2月28日的Nucleic Acids Reseach雜志上,。
同源重組（HR）是精確修復受損DNA的一種機制,，該過程利用來自同伴DNA的同源序列作為模板。這個過程包括將兩個DNA分子集合在一起,，搜索同源序列和交換DNA鏈,。
RecA家族蛋白是同源重組的中心重組酶,。這個家族的成員包括原核RecA,、古細菌RadA和真核Rad51和Dmc1,。它們在細胞增殖、基因組維護和遺傳多樣性中起到重要作用,，尤其是在高等真核生物中,。例如,，缺少Rad51的脊椎動物細胞會累積染色體缺口。Rad51和它的減數(shù)分裂特異性同系物Dmc1也是減數(shù)分裂過程必不可少的酶,。
自從研究人員發(fā)現(xiàn)RecA家族蛋白，研究人員就推測RecA和它的類似物只形成61右手螺旋細絲（每個螺旋由六個蛋白單體組成）,，然后水解ATP來促進同源重組和重組性的DNA修復,。然而始終存在一個爭議性的謎團——在鏈交換反應過程中,，ATP能量如何促進DNA的旋轉(zhuǎn),。
通過使用X射線晶體衍射和原子顯微鏡技術(shù),，王博士的研究組給出了這個問題的答案,。他們報道說,，古細菌Sulfolobus solfataricus的RadA蛋白還能夠自我聚合成一種31右手螺旋細絲（每個螺旋有3個單體）和一種43右手螺旋細絲（每個螺旋有4個單體）,。
另外的生物物理和生物化學分析揭示出RecA家族蛋白可能將ATP結(jié)合和水解過程以一種促進核蛋白順時針旋轉(zhuǎn)的方式來耦聯(lián)到DNA鏈的交換過程中,。尤其是61RadA螺旋細絲順時針旋轉(zhuǎn)變成31延伸的右手螺旋細絲,，然后轉(zhuǎn)化成43左手螺旋纖絲,。因此,，RadA蛋白中的所有DNA結(jié)合motifs一致運動來介導DNA結(jié)合、同源配對和鏈交換,。因此,，ATP能量不僅被用于DNA底物,，而且還被RecA家族蛋白纖絲所利用。這種新的模型挑戰(zhàn)了所有目前提出的假說,。
王惠鈞是臺灣大學化學系所畢業(yè),，美國伊利諾大學香檳分?；瘜W博士、麻省理工學院生物系博士后研究員到首席與資深研究員,，伊大香檳分校細胞及結(jié)構(gòu)生物學系教授,，中研院分子生物研究所學術(shù)咨詢委員,，歐洲生物化學學報編者,，臺大生化科學所教授,。
2000年當選中央研究院院士后,，回臺灣任中研院生物化學研究所特聘研究員兼所長至今，王惠鈞又陸續(xù)接任中華民國生物物理學會理事長,、臺灣生物化學及分子生物學會理事長,、臺灣蛋白質(zhì)體學會理事長,、世界蛋白質(zhì)體學會（HUPO）理事長,、亞太蛋白質(zhì)體學會（AOHUPO）理事,；他還曾獲美國生化及分生學會,、美國化學研究所,、美國科學促進協(xié)會等的名譽會員殊榮,。
王惠鈞一直致力提升臺灣的生化學術(shù)水平,，運用他生物結(jié)構(gòu)學研究專長,，展開生物結(jié)構(gòu)學與功能基因體學的探討，以了解重要的生物系統(tǒng)功能,，主要技術(shù)平臺有高效能的蛋白質(zhì)同步輻射結(jié)晶學與蛋白質(zhì)體學,，其它先進技術(shù)也可在需要時加入使用。
目前他正在努力四項領(lǐng)域,，結(jié)構(gòu)酶學是研究幾個具藥物發(fā)展潛力的酶群,，如蛋白質(zhì)分解酶、醣化酶,、磷酸水解酶,、prenyltransferases；蛋白質(zhì)-DNA與抗癌藥物-DNA交互作用是研究DNA結(jié)合蛋白和重要的抗癌藥物與DNA的交互作用,。
原始出處:
PLoS one
Received: July 20, 2007; Accepted: August 16, 2007; Published: September 12, 2007
Structural and Functional Analyses of Five Conserved Positively Charged Residues in the L1 and N-Terminal DNA Binding Motifs of Archaeal RadA Protein
Li-Tzu Chen1,3, Tzu-Ping Ko3, Yu-Wei Chang1,3, Kuei-An Lin3, Andrew H.-J. Wang1,2,3,4*, Ting-Fang Wang1,3*
1 Institute of Biochemical Sciences, National Taiwan University, Taipei, Taiwan, 2 Department of Life Sciences, National Taiwan University, Taipei, Taiwan, 3 Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan, 4 National Core Facility of High-Throughput Protein Crystallography, Academia Sinica, Taipei, Taiwan
RecA family proteins engage in an ATP-dependent DNA strand exchange reaction that includes a ssDNA nucleoprotein helical filament and a homologous dsDNA sequence. In spite of more than 20 years of efforts, the molecular mechanism of homology pairing and strand exchange is still not fully understood. Here we report a crystal structure of Sulfolobus solfataricus RadA overwound right-handed filament with three monomers per helical pitch. This structure reveals conformational details of the first ssDNA binding disordered loop (denoted L1 motif) and the dsDNA binding N-terminal domain (NTD). L1 and NTD together form an outwardly open palm structure on the outer surface of the helical filament. Inside this palm structure, five conserved basic amino acid residues (K27, K60, R117, R223 and R229) surround a 25 Å pocket that is wide enough to accommodate anionic ssDNA, dsDNA or both. Biochemical analyses demonstrate that these five positively charged residues are essential for DNA binding and for RadA-catalyzed D-loop formation. We suggest that the overwound right-handed RadA filament represents a functional conformation in the homology search and pairing reaction. A new structural model is proposed for the homologous interactions between a RadA-ssDNA nucleoprotein filament and its dsDNA target.
Received: July 20, 2007; Accepted: August 16, 2007; Published: September 12, 2007
Figure 1. Crystal packing and quaternary structures.
(A) SsoRadA protomers packed into three extended helical filaments. Chain A was located at the origin of the unit cell, whereas chains B and C were located one-third and two-third diagonal to the unit cell. (B) Side view of the SsoRadA right-handed helical filament crystal structure. The helical pitch of the filament is 98 Å. Each protomer is shown in a different color. The N-terminal domain (NTD), polymerization motif (PM), and central ATPase domain are indicated. (C) The Phe73 of the PM is buried in the hydrophobic pocket of the neighboring ATPase domain. Several hydrophobic residues that interact with the Phe73 side chain are indicated. The interactions result in the assembly of SsoRadA protomers into a filament. 2Fo–Fc electron density maps (contoured at 1.0 σ), corresponding to the PM are shown in orange.
全文鏈接：
