Theory and simulation of DNA repair enzymes; mechanism, structure and function Project Summary Human genome integrity depends on many processes to ensure the fidelity of the duplication of DNA.The efficiency of these processes is crucial since errors in DNA can often be key to disease onset. An important process to insure genome integrity is the repair of damaged DNA.[1] There are several types of DNA damage including (but not limited to): alkylation, oxidation, hydrolysis, adduct formation, base mismatch, among others. Alkylated DNA bases may be removed by two main routes: excision of the damaged base and activation of the base excision repair (BER) process, or direct dealkylation. The former route involves several enzymes involved in the BER cascade. The latter route may be performed by the AlkB family of enzymes. AlkB family enzymes are non-heme iron and ?-ketoglutarate dependent enzymes that perform an oxidative dealkylation of DNA. Some cancer treatments involve alkylating agents, and attempts have been made to enhance these therapies by inhibiting alkylating damage repair. Information gained from a detailed understanding of the structure and reaction mechanism of AlkB family proteins can aid in the development of inhibitors for these enzymes by providing useful information to develop transition state analogue inhibitors. One approach for this is via computational methods, including quantum mechanical/molecular mechanical (QM/MM) methods. Currently, most QM/MM implementations employ force fields that may not accurately describe the MM environment at close range, are not polarizable and lack methods to include long-range electrostatic effects. Our long-term goal is to understand the mechanism, structure and function of enzymes involved in DNA repair by means of computational simulations. To this end, the goals of GM108583 are: i) To study the structure/function/reactivity of AlkB family of enzymes by quantum mechanical/molecular mechanical (QM/MM), molecular dynamics (MD) and homology modeling. ii) To develop the first QM/MM program that interfaces a QM program with a two advanced force fields (GEM and AMOEBA) to accurately describe the MM environment; and to develop a novel method to introduce long-range electrostatic effects in QM/MM simulations. The detailed understanding of the structure, function and reaction mechanism of AlkB and its human homologues will provide insights into possible methods to inhibit these enzymes. In the current funding cycle we have already developed LICHEM [2] and pmemd.gem [3], collaborated in the development of TINKER?HP [4], and created QM/MM?LREC for long?range electrostatics in QM/MM [5, 6]. We reported new insights on ALKB enzymes including detailed understanding of the reaction mechanism of AlkB [7, 8]; confirmed an intra?molecular O2 tunnel in AlkB [9], and a homology model for ALKBH1 [10]. We used our HyDn?SNP?S method to uncover the first ever biomarker for prostate cancer on ALKBH7, and computationally predicted it?s effect on substrate binding, which was confirmed experimentally by our collaborators [11]. In addition we established collaborations with other experimental groups to investigate different DNA modification enzymes [12?15] and to improve force fields for computational simulations [16?20]. We are continuing our investigation of the reaction mechanism of ALKBH2 and ALKBH3 as well as expanding our investigation of enzymes of this family as well as other related enzymes based on our newly established collaborations.

Public Health Relevance

Theory and simulation of DNA repair enzymes; mechanism, structure and function Project Narrative The purpose of this proposal is to request an administrative supplement for the purchase of a single piece of equipment for data storage to enable the safe and efficient transfer and maintenance of the data generated by our research. The current setup for the equipment available in the Cisneros group is explained in detail in the equipment attachment. Briefly, the Cisneros group has a 36-node cluster that is located in the Center for Advanced Scientific Computing and Modeling cluster room in the 3rd floor of the Department of Chemistry at UNT (CASCaM). The Cisneros group also has three storage appliances, which are located in a separate server room that is located within the Cisneros group trainee suite in the 2nd floor of the Department of Chemistry at UNT (see facilities and other resources attachment). The Cisneros computer cluster has been our main compute tool for all of the latest projects mentioned above. Several of the above referenced projects have resulted and continue to generate several terabytes (TB) of data. The main issue with the current setup is that the network setup within the Department of Chemistry and cluster room at CASCaM/server room in the Cisneros office, do not allow our storage appliances to be connected directly to our cluster. This has resulted in a situation where we have to transfer several TB of data between our cluster and our storage servers. However, given the current network setup, the transfer speeds are extremely slow (less than 10 MB/s). These transfer speeds effectively have resulted in the participants having to wait some times over a week to transfer some of the generated data, and in a worst case scenario, some of transfers being interrupted. Based on the above issues, the principal goal of this request is to secure funds to purchase a storage server that will be installed and maintained within the CASCaM cluster room, and which will be connected directly to the Cisneros cluster. Since the racks in the cluster room are full, including the one that holds the Cisneros cluster, the quoted storage appliance includes a storage node (300 TB raw/239 TB RAIDed storage) with separate rack and required equipment (PDUs, etc), four high-speed (Infiniband) switches and a separate node for management/expansion. The total cost of the storage server is quoted at $199,038 (see attached quotes). The PI has secured a combined $32,500 in matching funds from the Department of Chemistry, College of Science and office of the Vice President for Research at the University of North Texas (see attached commitment letters). Thus, the total request for this supplement is $166,538. This storage solution will provide a much needed reprieve for storage in our group and will allow us to continue and expand our proposed simulations on the ALKB family enzymes, as well as expand our collaborations with the other groups on DNA related enzymes.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
Project #
Application #
Study Section
Program Officer
Lyster, Peter
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
University of North Texas
Schools of Arts and Sciences
United States
Zip Code
Silvestrov, Pavel; Cisneros, G Andrés (2018) Insights into conformational changes in AlkD bound to DNA with a yatakemycin adduct from computational simulations. Theor Chem Acc 137:
Gahlon, Hailey L; Walker, Alice R; Cisneros, G Andrés et al. (2018) Reduced structural flexibility for an exonuclease deficient DNA polymerase III mutant. Phys Chem Chem Phys 20:26892-26902
Silvestrov, Pavel; Maier, Sarah J; Fang, Michelle et al. (2018) DNArCdb: A database of cancer biomarkers in DNA repair genes that includes variants related to multiple cancer phenotypes. DNA Repair (Amst) 70:10-17
Gökcan, Hatice; Kratz, Eric; Darden, Thomas A et al. (2018) QM/MM Simulations with the Gaussian Electrostatic Model: A Density-based Polarizable Potential. J Phys Chem Lett 9:3062-3067
Antczak, Nicole M; Walker, Alice R; Stern, Hannah R et al. (2018) Characterization of Nine Cancer-Associated Variants in Human DNA Polymerase ?. Chem Res Toxicol 31:697-711
Torabifard, Hedieh; Cisneros, G Andrés (2017) Computational investigation of O2 diffusion through an intra-molecular tunnel in AlkB; influence of polarization on O2 transport. Chem Sci 8:6230-6238
Walker, Alice R; Cisneros, G Andrés (2017) Computational Simulations of DNA Polymerases: Detailed Insights on Structure/Function/Mechanism from Native Proteins to Cancer Variants. Chem Res Toxicol 30:1922-1935
Liu, Monica Yun; Torabifard, Hedieh; Crawford, Daniel J et al. (2017) Mutations along a TET2 active site scaffold stall oxidation at 5-hydroxymethylcytosine. Nat Chem Biol 13:181-187
Walker, Alice R; Silvestrov, Pavel; Müller, Tina A et al. (2017) ALKBH7 Variant Related to Prostate Cancer Exhibits Altered Substrate Binding. PLoS Comput Biol 13:e1005345
Kratz, Eric G; Walker, Alice R; Lagardère, Louis et al. (2016) LICHEM: A QM/MM program for simulations with multipolar and polarizable force fields. J Comput Chem 37:1019-29

Showing the most recent 10 out of 16 publications