COMPARISON OF MOLECULAR DYNAMICS SIMULATIONS OF DIHYDROFOLATE REDUCTASE BOUND T

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 机会性真菌感染在过去几十年中显着增加，产生了对新的抗真菌疗法的需求(1-3)。目前，我们正在进行一项针对二氢叶酸还原酶(DHFR)的药物设计计划，目标是使用炔丙基连接的抑制剂。DHFR是一个公认的药物靶点，对所有生物体的DNA和蛋白质合成都是必不可少的(4-7)。白念珠菌二氢叶酸还原酶(CaDHFR)(8)、光滑念珠菌二氢叶酸还原酶(CgDHFR)和人DHFR(9)的模型已经被创建，以对接内部合成的炔丙基连接抗叶酸的文库。为了使对接模型与经验确定的抑制结果相关联，有必要将关键活性部位残基视为灵活的。有限分子动力学(MD)模拟被用来允许配体半径3.5内的残基移动。最重要的是，活性位点两侧的环残基(CaDHFR和CgDHFR：Thr 58-Phe 66，HuDHFR：Thr 56-ASN)的灵活性被发现对对接精度至关重要。沿着MD轨道拍摄了快照，以创建用于对接的结构集合。灵活性提高了对接结果的准确性(10)，并与新确定的与炔丙基连接的反叶酸结合的CaDHFR和CgDHFR的晶体结构很好地相关。这些环残基和炔丙基连接的配体之间的相互作用对真菌的效力和人类的选择性都起着关键作用。当真菌DHFR和huDHFR的结构比对时，环残基在离叶酸结合部位不同的距离处移位，因此在活性部位占据不同的体积，其中人的体积最小。有人认为，增加配体的大小，从而在huDHFR结合部位产生不利的空间干扰，将增加对真菌DHFR的选择性。然而，这种设计策略取得的成功有限。结合位点两侧的环的灵活性质允许huDHFR结合位点容纳被设计为对真菌DHFR具有选择性的配体。为了了解这些环残基在效力和选择性方面所起的作用，我们将在更长的时间尺度上进行更全面的MD模拟，包括包括溶剂在内的整个体系。模拟结果将通过监测瞬时和永久氢键、比较不同物种中的环位移以及使用不同的配体支架来分析。Desmond(D.E.Shaw)是一个专门为分子动力学模拟设计的高度可并行化的程序，将用于执行所有模拟。因此，请求对安装了Desmond的四个资源进行分配。1.M.B.Edmond，S.E.Wallace，D.K.McClish，M.A.Pfall，R.N.Jones和R.P.Wenzel，Clin。感染。DIS，1999，29,239-244.2.M.A.Pfall和D.J.Diekema，J.Clin.《微生物》，2004，42,4419-4431。3.R.A.Hajjeh，A.N.Sofair，L.H.Harison，G.M.Lyon，B.A.阿尔辛顿-斯卡格斯，S.A.米孜，M.Phelan，J.摩根，W.Lee-Yang，M.A.Ciblak，L.E.Benjamin，L.T.Sanza，S.Huie，S.F.Yeo，M.E.Brandt和D.W.Warnock，J.Clin。《微生物》，2004，42,1519-1527。4.G·F·弗莱明和R.L.希尔斯基，塞明。Oncol.，1992，19,707-719.5.B.Roth，B.S.Ruckman，R.Ferone，D.P.Baccanari，J.N.Champness和R.M.Hyde，J.化学，1987，30,348-356。6.A.J.Salter，Rev.Infect。DIS，1982，4,196-236.7.C.Plowe，J.Kublin，D.Kamwendo，R.Mukadam，C.P.，M.Molyneux，T.Taylor和E.Terrie，Brit.地中海医院。J.，2004,545-548。8.J.L.Paulsen，J.Liu，D.B.Bolstad，A.E.Smith，N.D.Priestley，D.L.Wright和A.C.Anderson，Bioorg。地中海医院。化学，2009，17,4866-4872。9.O.Algul，J.L.Paulsen和A.C.Anderson，J.Mol。图表。型号，2011，29,608-613。10.J.L.Paulsen和A.C.Anderson，J.信息型号，2009，49,2813-2819。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Opportunistic fungal infections have increased significantly over the last few decades creating a need for new antifungal therapies (1-3). Presently, we have a drug design program underway aimed at targeting Dihydrofolate reductase (DHFR) with propargyl-linked inhibitors. DHFR is a well established drug target that is essential for DNA and protein synthesis in all organisms (4-7). Models of Candida albicans dihydrofolate reductase (CaDHFR) (8), Candida glabrata dihydrofolate reductase (CgDHFR) and human DHFR (9) have been created to dock an in-house library of synthetically generated propargyl-linked antifolates. In order for docking models to correlate with empirically determined inhibition results it was necessary to treat key active site residues as flexible. Limited molecular dynamics (MD) simulations were used to allow residues within 3.5 radius of the ligand to move. Most importantly, the flexibility of loop residues (CaDHFR and CgDHFR: Thr 58-Phe 66, HuDHFR: Thr 56-Asn 64) flanking the active site was found to be critical to docking accuracy. Snapshots were taken along the MD trajectory to create an ensemble of structures for docking. Flexibility increased the accuracy of docking results (10) and correlates well with newly determined crystallographic structures of CaDHFR and CgDHFR bound to propargyl-linked antifolates. The interactions between these loop residues and the propargyl-linked ligands play a key role in both fungal potency and human selectivity. When a structural alignment of fungal DHFR is compared with huDHFR the loop residues are displaced at different distances from the folate binding site and therefore, occupy different volumes in the active site with human having the smallest volume. It was thought that increasing the size of the ligand, thus creating unfavorable steric interference in the huDHFR binding site would increase selectivity for fungal DHFR. However, this design strategy has met with limited success. The flexible nature of the loop flanking the binding site allows the huDHFR binding site to accommodate ligands that were designed to be selective for fungal DHFR. In order to understand the role these loop residues play in potency and selectivity we will perform more comprehensive MD simulations at longer time scales that will encompass the entire system including solvent. The resultant simulations will be analyzed by monitoring transient and permanent hydrogen bonds, comparing loop displacement in different species and using different ligand scaffolds. Desmond (D.E. Shaw) which is a highly parallelizable program specifically designed for molecular dynamics simulations will be used to perform all simulations. Therefore, allocations on four resources that have Desmond installed are requested. 1. M. B. Edmond, S. E. Wallace, D. K. McClish, M. A. Pfaller, R. N. Jones and R. P. Wenzel, Clin. Infect. Dis., 1999, 29, 239-244. 2. M. A. Pfaller and D. J. Diekema, J. Clin. Microbiol., 2004, 42, 4419-4431. 3. R. A. Hajjeh, A. N. Sofair, L. H. Harrison, G. M. Lyon, B. A. Arthington-Skaggs, S. A. Mirza, M. Phelan, J. Morgan, W. Lee-Yang, M. A. Ciblak, L. E. Benjamin, L. T. Sanza, S. Huie, S. F. Yeo, M. E. Brandt and D. W. Warnock, J. Clin. Microbiol., 2004, 42, 1519-1527. 4. G. F. Fleming and R. L. Schilsky, Semin. Oncol., 1992, 19, 707-719. 5. B. Roth, B. S. Rauckman, R. Ferone, D. P. Baccanari, J. N. Champness and R. M. Hyde, J. Med. Chem., 1987, 30, 348-356. 6. A. J. Salter, Rev. Infect. Dis., 1982, 4, 196-236. 7. C. Plowe, J. Kublin, D. Kamwendo, R. Mukadam, C. P, M. Molyneux, T. Taylor and E. Terrie, Brit. Med. J., 2004, 545-548. 8. J. L. Paulsen, J. Liu, D. B. Bolstad, A. E. Smith, N. D. Priestley, D. L. Wright and A. C. Anderson, Bioorg. Med. Chem., 2009, 17, 4866-4872. 9. O. Algul, J. L. Paulsen and A. C. Anderson, J. Mol. Graph. Model., 2011, 29, 608-613. 10. J. L. Paulsen and A. C. Anderson, J. Chem. Inf. Model., 2009, 49, 2813-2819.