Single molecule studies of the dual-helicase activity of transcription factor TFIIH

项目总结/摘要 候选人寻求获得额外的研究和职业发展培训，通过一个计划， 以弥补她现有专业知识的不足该计划将利用她在以下方面的现有技能和知识： 研究一个新的系统，她将带着这个系统进入她职业生涯的下一个阶段。她将使用 该奖项的指导阶段，以建立在仪器设计，人员管理和手稿的技能 发展和出版。这项培训将使她获得过渡所需的额外技能 独立并建立一个成功的研究计划。 拟议的研究将使用尖端的单分子和计算技术来研究 DNA解旋复合物TFIIH的酶活性。TFIIH为转录和DNA打开DNA 修复，但利用具有不同机制的两个单独的解旋酶结构域来这样做。该提案采用了 结合单分子磁镊研究和分子动力学模拟： 表征TFIIH的单个解旋酶结构域的活性（目标1）;研究两者的影响 在完整的TFIIH复合物中，解旋酶相互作用（目的2）;并探索 两个解旋酶和导致一个比另一个占优势的DNA结构方面（目的3）。 该研究项目的完成将有助于转录和核苷酸切除领域 通过阐明TFIIH连接这两种途径的方式来修复。拟议研究的结果 将为进一步研究TFIIH活动及其影响因素提供基础，并形成基础 候选人的第一个R 01提案 K22奖将为候选人提供机会，通过额外的研究开发新的研究技能。 在指导阶段的培训，这将是由NHLBI校内研究司资助。这些 技能包括构造组合的磁性镊子/单分子荧光显微镜， 单分子实验的DNA底物设计策略和数据分析方法。 在该奖项的指导阶段，她还将参加旨在传授 管理和教学技能，并监督一个学士学位后的实习生。她将参加研讨会， 手稿准备和建立在她目前的出版记录。培训计划还包括职业 发展活动，如实践工作会谈，模拟面试，并授予写作讲习班。这些 活动将为她提供宝贵的经验和技能，她将借鉴作为一个独立的 调查员

Project Summary/Abstract The candidate seeks to acquire additional research and career development training through a plan designed to address deficiencies in her current expertise. The plan will build on her existing skills and knowledge in the study of a new system which she will carry with her into the next phase of her career. She will use the mentored phase of this award to build skills in instrument design, personnel management, and manuscript development and publication. This training will enable her to acquire the additional skills necessary to transition to independence and establish a successful research program. The proposed research will use cutting edge single-molecule and computational techniques to study the enzymatic activity of the DNA-unwinding complex TFIIH. TFIIH opens DNA for both transcription and DNA repair, but utilizes two separate helicase domains with distinct mechanisms to do so. The proposal employs a combination of single-molecule magnetic tweezers studies and molecular dynamics simulations to: characterize the activity of the individual helicase domains of TFIIH (Aim 1); investigate the impact of the two helicases on each other in the full TFIIH complex (Aim 2); and explore the differences in DNA preference of the two helicases and the aspects of DNA structure that lead to dominance of one over the other (Aim 3). Completion of this research project will contribute to the fields of both transcription and nucleotide excision repair by elucidating the ways in which TFIIH links these two pathways. The results of the proposed research will provide a foundation for further studies of TFIIH activity and the factors that influence it, and form the basis of the candidate's first R01 proposal. The K22 award will provide the candidate with the opportunity to develop new research skills through additional training during the mentored phase, which will be funded by the NHLBI Division of Intramural Research. These skills include construction of a combined magnetic tweezers/single molecule fluorescence microscope, strategies for the design of DNA substrates for single-molecule experiments, and data analysis methods. During the mentored phase of the award she will also attend courses and seminars designed to impart management and teaching skills and supervise a post-baccalaureate trainee. She will attend workshops on manuscript preparation and build on her current publication record. The training plan also includes career development activities such as practice job talks, mock interviews, and grant writing workshops. These activities will provide her with invaluable experience and skills that she will draw on as an independent investigator.