J Biol Chem:董梦秋实验室与合作者成功开发了研究蛋白质动态的新方法

2017-04-20 佚名 北京生命科学研究所

日前,北京生命科学研究所董梦秋实验室与武汉物理与数学研究所唐淳实验室合作在《The Journal of Biological Chemistry》发表题为“Modeling protein excited-state structures from "over-length" chemical cross-links”的研究论文。作者成功探索了利用化学交联质谱(chemical cross-li

日前,北京生命科学研究所董梦秋实验室与武汉物理与数学研究所唐淳实验室合作在《The Journal of Biological Chemistry》发表题为“Modeling protein excited-state structures from "over-length" chemical cross-links”的研究论文。作者成功探索了利用化学交联质谱(chemical cross-linking of proteins coupled with mass spectrometry, or CXMS)和结构计算来研究蛋白质动态的新途径,并发布了通用流程。文章的艺术配图被选为当期JBC封面。

到目前为止,结构生物学研究主要关注蛋白质的基态结构,但是蛋白质行使功能往往需要在基态和激发态之间转变构象。解析蛋白质的激发态构象和动态结构对于X射线晶体学和电镜等传统结构生物学方法是一大难题。核磁共振(NMR)虽然适合研究蛋白质在溶液中的多种构象,但是仅限于50 kDa以下的蛋白,而且需要大量的高度纯化的蛋白。

董梦秋实验室一直从事化学交联质谱技术CXMS的开发和应用(Yang B, Nature Methods 2012; Lu S, Nature Methods 2015; Tan D, eLife 2016)。蛋白样品中两个氨基酸之间发生交联反应,必须满足交联剂的化学特异性要求(如氨基特异性)以及空间距离小于或等于交联剂臂长的要求。因此,由CXMS获得的交联位点对提供了可用于结构计算的距离约束信息。作为结构生物学的新工具,特别是单分子电镜技术的辅助工具,CXMS在很多大型蛋白质复合体的结构解析中做出了贡献。但是,这个技术还存在一些问题,其中最令人困扰的是:即便用最严苛的筛选标准,总有小部分交联位点对在晶体结构(一般是基态构象)中的距离大于交联剂臂长("over-length" chemical cross-links)。对于此类数据,大部分实验室选择丢弃或者放松距离约束(如果结构未知)。本文两个实验室一年前合作发现,分子间的高可信度过长交联反映了蛋白-蛋白间的弱(或瞬时)相互作用(Gong Z, Biophysics Reports 2015)。本文重点研究了蛋白内不同结构域之间的相对运动。实验结果表明,分子内的高可信度过长交联来源于蛋白质的激发态构象;它们不是制造麻烦的问题数据,而是“众里寻他千百度”的反映蛋白质动态的珍贵信息。作者通过交联数据和结构计算发现,结合了钙离子但没有结合配体肽段或蛋白的钙调蛋白(Ca2+-CaM) 除了采取哑铃状的开放态构象(基态),还有一种闭合态构象,非常接近之前用NMR手段观察到的激发态构象。这种闭合态构象与结合了各种配体的Ca2+-CaM的晶体结构或NMR结构也非常相似,说明配体肽段或蛋白的结合稳定了原已存在的Ca2+-CaM的闭合态构象。本文还用同样方法研究了细菌磷酸转移系统中的蛋白EIN和谷氨酰胺结合蛋白QBP,发现它们均含有基态构象之外的一种激发态构象。

本文建立了一套基于交联数据的结构模拟计算方法,内容包括:用15N同位素标记的蛋白与相对应的天然同位素标记的蛋白等量混合后进行交联质谱分析,从中得到高可信度的分子内交联数据,然后采用专门开发的DynaXL软件进行构象模拟计算。DynaXL包含结构域识别、模拟交联反应、分子动力学模拟等多个模块。它首先尝试用基态构象模拟交联反应,如果不能满足全部交联约束,则添加一个构象,直到所有交联约束都得以满足。同类其它算法只考虑单一构象,试图让单个构象满足所有交联数据;当蛋白质在溶液中存在多种构象时,容易造成过拟合,得到错误结构。

已发表的NMR研究表明,Ca2+-CaM的激发态构象在溶液中所占的比例小于5%;EIN和QBP的激发态构象很可能由于丰度过低未能被NMR检测到。这说明CXMS-DynaXL方法可以灵敏地检测到溶液中的低丰度构象,灵敏度达到甚至超过NMR,且不受蛋白大小限制,可以广泛应用于蛋白质动态变化研究。

董梦秋实验室的丁曰和博士以及唐淳实验室的龚洲博士为本文共同第一作者。为本工作做出贡献的还有唐淳实验室的董旭和刘侃,浙江大学的刘主以及中科院计算所的贺思敏和刘超。董梦秋和唐淳为本文共同通讯作者。

原始出处:

Yue-He Ding, Zhou Gong, Xu Dong,et al.Modeling protein excited-state structures from "over-length" chemical cross-links.J Biol Chem. 2017 Jan 27

版权声明:
本网站所有内容来源注明为“梅斯医学”或“MedSci原创”的文字、图片和音视频资料,版权均属于梅斯医学所有。非经授权,任何媒体、网站或个人不得转载,授权转载时须注明来源为“梅斯医学”。其它来源的文章系转载文章,或“梅斯号”自媒体发布的文章,仅系出于传递更多信息之目的,本站仅负责审核内容合规,其内容不代表本站立场,本站不负责内容的准确性和版权。如果存在侵权、或不希望被转载的媒体或个人可与我们联系,我们将立即进行删除处理。
在此留言
评论区 (5)
#插入话题
  1. [GetPortalCommentsPageByObjectIdResponse(id=1917444, encodeId=5c4b191e444fc, content=<a href='/topic/show?id=4f26341994' target=_blank style='color:#2F92EE;'>#Bio#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=26, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3419, encryptionId=4f26341994, topicName=Bio)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=12de429, createdName=sunylz, createdTime=Sun Feb 18 19:58:00 CST 2018, time=2018-02-18, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1767098, encodeId=3a721e6709832, content=<a href='/topic/show?id=64f5343e2f' target=_blank style='color:#2F92EE;'>#Biol#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=29, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3437, encryptionId=64f5343e2f, topicName=Biol)], attachment=null, authenticateStatus=null, createdAvatar=null, createdBy=446937890434, createdName=12498ebem31暂无昵称, createdTime=Sun Aug 13 14:58:00 CST 2017, time=2017-08-13, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1765615, encodeId=64e81e6561569, content=<a href='/topic/show?id=252ae1803a3' target=_blank style='color:#2F92EE;'>#白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=71803, encryptionId=252ae1803a3, topicName=白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=778d36, createdName=柳叶一刀, createdTime=Sun Oct 22 16:58:00 CST 2017, time=2017-10-22, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1903214, encodeId=56421903214e5, content=<a href='/topic/show?id=e1b0583884b' target=_blank style='color:#2F92EE;'>#新方法#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=58388, encryptionId=e1b0583884b, topicName=新方法)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c22f171, createdName=kord1982, createdTime=Mon Sep 11 14:58:00 CST 2017, time=2017-09-11, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1296956, encodeId=9654129695697, content=<a href='/topic/show?id=158288e017c' target=_blank style='color:#2F92EE;'>#蛋白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=0, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=88701, encryptionId=158288e017c, topicName=蛋白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c689267, createdName=cmj8wellington, createdTime=Sat Apr 22 06:58:00 CST 2017, time=2017-04-22, status=1, ipAttribution=)]
    2018-02-18 sunylz
  2. [GetPortalCommentsPageByObjectIdResponse(id=1917444, encodeId=5c4b191e444fc, content=<a href='/topic/show?id=4f26341994' target=_blank style='color:#2F92EE;'>#Bio#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=26, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3419, encryptionId=4f26341994, topicName=Bio)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=12de429, createdName=sunylz, createdTime=Sun Feb 18 19:58:00 CST 2018, time=2018-02-18, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1767098, encodeId=3a721e6709832, content=<a href='/topic/show?id=64f5343e2f' target=_blank style='color:#2F92EE;'>#Biol#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=29, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3437, encryptionId=64f5343e2f, topicName=Biol)], attachment=null, authenticateStatus=null, createdAvatar=null, createdBy=446937890434, createdName=12498ebem31暂无昵称, createdTime=Sun Aug 13 14:58:00 CST 2017, time=2017-08-13, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1765615, encodeId=64e81e6561569, content=<a href='/topic/show?id=252ae1803a3' target=_blank style='color:#2F92EE;'>#白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=71803, encryptionId=252ae1803a3, topicName=白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=778d36, createdName=柳叶一刀, createdTime=Sun Oct 22 16:58:00 CST 2017, time=2017-10-22, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1903214, encodeId=56421903214e5, content=<a href='/topic/show?id=e1b0583884b' target=_blank style='color:#2F92EE;'>#新方法#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=58388, encryptionId=e1b0583884b, topicName=新方法)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c22f171, createdName=kord1982, createdTime=Mon Sep 11 14:58:00 CST 2017, time=2017-09-11, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1296956, encodeId=9654129695697, content=<a href='/topic/show?id=158288e017c' target=_blank style='color:#2F92EE;'>#蛋白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=0, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=88701, encryptionId=158288e017c, topicName=蛋白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c689267, createdName=cmj8wellington, createdTime=Sat Apr 22 06:58:00 CST 2017, time=2017-04-22, status=1, ipAttribution=)]
  3. [GetPortalCommentsPageByObjectIdResponse(id=1917444, encodeId=5c4b191e444fc, content=<a href='/topic/show?id=4f26341994' target=_blank style='color:#2F92EE;'>#Bio#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=26, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3419, encryptionId=4f26341994, topicName=Bio)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=12de429, createdName=sunylz, createdTime=Sun Feb 18 19:58:00 CST 2018, time=2018-02-18, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1767098, encodeId=3a721e6709832, content=<a href='/topic/show?id=64f5343e2f' target=_blank style='color:#2F92EE;'>#Biol#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=29, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3437, encryptionId=64f5343e2f, topicName=Biol)], attachment=null, authenticateStatus=null, createdAvatar=null, createdBy=446937890434, createdName=12498ebem31暂无昵称, createdTime=Sun Aug 13 14:58:00 CST 2017, time=2017-08-13, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1765615, encodeId=64e81e6561569, content=<a href='/topic/show?id=252ae1803a3' target=_blank style='color:#2F92EE;'>#白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=71803, encryptionId=252ae1803a3, topicName=白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=778d36, createdName=柳叶一刀, createdTime=Sun Oct 22 16:58:00 CST 2017, time=2017-10-22, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1903214, encodeId=56421903214e5, content=<a href='/topic/show?id=e1b0583884b' target=_blank style='color:#2F92EE;'>#新方法#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=58388, encryptionId=e1b0583884b, topicName=新方法)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c22f171, createdName=kord1982, createdTime=Mon Sep 11 14:58:00 CST 2017, time=2017-09-11, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1296956, encodeId=9654129695697, content=<a href='/topic/show?id=158288e017c' target=_blank style='color:#2F92EE;'>#蛋白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=0, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=88701, encryptionId=158288e017c, topicName=蛋白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c689267, createdName=cmj8wellington, createdTime=Sat Apr 22 06:58:00 CST 2017, time=2017-04-22, status=1, ipAttribution=)]
  4. [GetPortalCommentsPageByObjectIdResponse(id=1917444, encodeId=5c4b191e444fc, content=<a href='/topic/show?id=4f26341994' target=_blank style='color:#2F92EE;'>#Bio#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=26, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3419, encryptionId=4f26341994, topicName=Bio)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=12de429, createdName=sunylz, createdTime=Sun Feb 18 19:58:00 CST 2018, time=2018-02-18, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1767098, encodeId=3a721e6709832, content=<a href='/topic/show?id=64f5343e2f' target=_blank style='color:#2F92EE;'>#Biol#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=29, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3437, encryptionId=64f5343e2f, topicName=Biol)], attachment=null, authenticateStatus=null, createdAvatar=null, createdBy=446937890434, createdName=12498ebem31暂无昵称, createdTime=Sun Aug 13 14:58:00 CST 2017, time=2017-08-13, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1765615, encodeId=64e81e6561569, content=<a href='/topic/show?id=252ae1803a3' target=_blank style='color:#2F92EE;'>#白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=71803, encryptionId=252ae1803a3, topicName=白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=778d36, createdName=柳叶一刀, createdTime=Sun Oct 22 16:58:00 CST 2017, time=2017-10-22, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1903214, encodeId=56421903214e5, content=<a href='/topic/show?id=e1b0583884b' target=_blank style='color:#2F92EE;'>#新方法#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=58388, encryptionId=e1b0583884b, topicName=新方法)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c22f171, createdName=kord1982, createdTime=Mon Sep 11 14:58:00 CST 2017, time=2017-09-11, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1296956, encodeId=9654129695697, content=<a href='/topic/show?id=158288e017c' target=_blank style='color:#2F92EE;'>#蛋白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=0, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=88701, encryptionId=158288e017c, topicName=蛋白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c689267, createdName=cmj8wellington, createdTime=Sat Apr 22 06:58:00 CST 2017, time=2017-04-22, status=1, ipAttribution=)]
  5. [GetPortalCommentsPageByObjectIdResponse(id=1917444, encodeId=5c4b191e444fc, content=<a href='/topic/show?id=4f26341994' target=_blank style='color:#2F92EE;'>#Bio#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=26, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3419, encryptionId=4f26341994, topicName=Bio)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=12de429, createdName=sunylz, createdTime=Sun Feb 18 19:58:00 CST 2018, time=2018-02-18, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1767098, encodeId=3a721e6709832, content=<a href='/topic/show?id=64f5343e2f' target=_blank style='color:#2F92EE;'>#Biol#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=29, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=3437, encryptionId=64f5343e2f, topicName=Biol)], attachment=null, authenticateStatus=null, createdAvatar=null, createdBy=446937890434, createdName=12498ebem31暂无昵称, createdTime=Sun Aug 13 14:58:00 CST 2017, time=2017-08-13, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1765615, encodeId=64e81e6561569, content=<a href='/topic/show?id=252ae1803a3' target=_blank style='color:#2F92EE;'>#白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=71803, encryptionId=252ae1803a3, topicName=白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=778d36, createdName=柳叶一刀, createdTime=Sun Oct 22 16:58:00 CST 2017, time=2017-10-22, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1903214, encodeId=56421903214e5, content=<a href='/topic/show?id=e1b0583884b' target=_blank style='color:#2F92EE;'>#新方法#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=27, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=58388, encryptionId=e1b0583884b, topicName=新方法)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c22f171, createdName=kord1982, createdTime=Mon Sep 11 14:58:00 CST 2017, time=2017-09-11, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1296956, encodeId=9654129695697, content=<a href='/topic/show?id=158288e017c' target=_blank style='color:#2F92EE;'>#蛋白质#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=0, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=88701, encryptionId=158288e017c, topicName=蛋白质)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c689267, createdName=cmj8wellington, createdTime=Sat Apr 22 06:58:00 CST 2017, time=2017-04-22, status=1, ipAttribution=)]