O2-1

REstretto: An Efficient Protein-Ligand Docking Tool
based on a Fragment Reuse Strategy
Keisuke Yanagisawa 1

yanagisawa@c.titech.ac.jp

Rikuto Kubota 1,2

kubota@bi.c.titech.ac.jp

Masahito Ohue 1

ohue@c.titech.ac.jp

12

Yasushi Yoshikawa 1

y_yoshikawa@bi.c.titech.ac.jp

Yutaka Akiyama 1akiyama@c.titech.ac.jp

Department of Computer Science, School of Computing, Tokyo Institute of Technology,
2-12-1 Ookayama, Meguro-ku, Tokyo, 152-8550, Japan
AIST-Tokyo Tech Real World Big-Data Computation Open Innovation Laboratory(RWBC-OIL), National Institute of Advanced Industrial Science and Technology,
Tsukuba, Ibaraki 305-8560, Japan

Keywords: Protein-ligand docking, Fragment commonality, Software implementation
Virtual screening is a commonly used process to search for feasible drug candidates from a huge
number of compounds during the early stages of drug design. As the compound database continues
to expand to billions of entries or more, there remains an urgent used to accelerate the process of
docking calculation. Reuse of calculation results is a possible way to accelerate the process.
In this study, we first propose novel virtual screening-oriented docking strategy by combining
three factors, namely, compound decomposition, simplified fragment grid storing k-best scores, andflexibility consideration with pre-generated conformers. Candidate compounds contain many
common fragments (chemical substructures). Thus, the calculation results of these common
fragments can be reused among them.
As a proof-of-concept of the aforementioned strategies, we also conducted the development ofREstretto, a tool that implements the three factors to enable the reuse of calculation results. We
demonstrated that the speed and accuracy of REstretto were comparable to those of AutoDock Vina,a well-known free docking tool. The implementation of REstretto has much room for further
performance improvement, and therefore, the results dhow the feasibility of the strategy.
The code is available under an MIT license at https://github.com/akiyamalab/restretto.
[1] Yanagisawa K.; Kubota R.; Yoshikawa Y.; Ohue M.; Akiyama Y. Effective Protein-Ligand
Docking Strategy via Fragment Reuse and a Proof-of-Concept Implementation. ACS Omega,
doi: 10.1021/acsomega.2c03470. (Epub ahead of print)

O2-2

Binding specificity of phosphopeptide recognition domainanalyzed by gREST simulation

12

Suyong Re 1

Kenji Mizuguchi 1,2

suyongre@nibiohn.go.jp

kenji@protein.osaka-u.ac.jp

Artificial Intelligence Center for Health and Biomedical Research, National Institutes ofBiomedical Innovation, Health, and Nutrition 7-6-8, Saito-Asagi, Ibaraki, Osaka, 567-0085,
Japan
Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan

Keywords: Protein-peptide binding, Molecular dynamics simulation, generalized
replica-exchange with solute tempering
Protein-protein interactions play an important role in intracellular signal transduction. Protein
phosphorylation is common in modulating interacting partners to induce signals of interest. Thus,
the binding specificity of phosphopeptide recognition domains is an important subject forunderstanding cellular function and for therapeutic purpose. Peptide array techniques andavailable three-dimensional structures of complexes provide a powerful means to determine thebinding specificity of phosphopeptide recognition domains. However, the underlying recognition
mechanisms remain elusive due to their dynamic and complex nature, making the development ofreliable predictive models difficult. Here, we apply the generalized replica-exchange with solutetempering method, gREST [1] implemented in GENESIS [2], which is a generalization ofREST/REST2 [3], to analyze the binding mechanism of phosphopeptides by Src homology 2 (SH2)domain. By selecting the phosphopeptide and the binding site residues as the solute region, theprotein-peptide interactions are effectively weakened, and many of the 16 replicas showed thebinding/unbinding events, which is not possible using conventional molecular dynamics
simulations. Details of binding mechanism and possible determinants of specificity will be
discussed.
[1] Kamiya, M.; Sugita, Y. J. Chem. Phys. 2018, 149, 072304.
[2] Jung, J.; Mori, T. et al., WIREs Comput. Mol. Science. 2015, 5, 310-323. Kobayashi, C.; Jung,J. et al., J. Comput. Chem. 2017, 38, 2193-2206.
[3] Wang, L. et al. J. Phys. Chem. B 2011, 115, 9431–9438. Moors, S. L. C. et al. J. Chem. Theory
Comput. 2011, 7, 231–237. Terakawa, T. et al. J. Comput. Chem. 2011, 32, 1228–1234.

O2-3

Dynamical interaction analysis of Remdesivir with SARS-CoV-2
RNA-dependent RNA polymerase by MD and FMO calculations
Shuhei Miyakawa1 Koji Okuwaki1s181232@hoshi.ac.jp

Yuma Handa
d2002@hoshi.ac.jp

1,3

k-okuwaki@hoshi.ac.jp

Takayuki Furuishi1 Etsuo Yonemochi1

t-furuishi@hoshi.ac.jp

e-yonemochi@hoshi.ac.jp

Koichiro Kato2 Kaori Fukuzawa1,3
kato.koichiro.957@m.kyushu-u.ac.jp fukuzawa-k@phs.osaka-u.ac.jp

123

Hoshi Univ., 2-4-41 Ebara, Shinagawa-ku, Tokyo, 142-8501, Japan
Kyushu Univ., 744 Motoka, Nishi-ku, Fukuoka, 819-0395, Japan
Osaka Univ., 1-6 Yamadaoka, Suita-shi, Osaka, 565-0871, Japan

Keywords: SARS-CoV-2, RNA dependent RNA polymerase (RdRp), Molecular Dynamics,
Fragment Molecular Orbital Method (FMO)
Remdesivir (Rem), one of the COVID-19 therapeutics, is incorporated into the RNA-dependent RNA
polymerase (RdRp) of the SARS-CoV-2 virus in competition with Adenine and inhibits RNA elongation whenlocated at the position -3 on the RNA recognized by RdRp. In this study, we aim to investigate the detailed
molecular mechanism of Rem.
Based on the cryo-EM structure of the RdRp-RNA-Rem complex (PDBID:7BV2), seven structures weregenerated: five structures in which bases at the positions +1 to -3 of the elongated strand in the RNA duplex
were replaced with Rem/Adenine (Rem+1 ～Rem-3, and A-3), a structure in which a cyano group was deleted
from Rem in Rem-3 (Rem’-3), and a structure in which Lys593 in Rem-3 was mutated to Ala (Rem[K593A]-3).
Then molecular dynamics (MD) calculations using the Amber20 program were performed for 50 ns for eachstructure. Finally, fragment molecular orbital (FMO) calculations were performed on a total of 70 structures
extracted from the MD trajectories using the ABINIT-MP program on the supercomputer Fugaku (hp220143)
to analyze the molecular interactions around Rem.
The inter-fragment interaction energy (IFIE) between the complementary strands in the RNA duplex wasexamined for Rem+1 to Rem-3, and it was confirmed that the hydrogen bonds of base pairs from the position+1 to -2 were disrupted only when Rem was located at the position -3. In the interaction between Rem at the 3 position and the surrounding residues, the cyano group of Rem acquired a cation-π interaction with theLys593. Therefore, we compared the average distance between Rem/Adenine at the -3 position and the Lys593,
and found that Rem maintained an interaction with the Lys593 at a distance 0.7 Å shorter than that of Adenine.
We compared the IFIE between Rem/Adenine at the -3 position and the Lys593, and found that Rem interactedwith the Lys593 about 16.8 kcal/mol more stable than Adenine. The structure of RNA duplex was maintainedin Rem’-3 and Rem[K593A]-3.
These results suggest that Rem affects downstream RNA structure by interacting with the Lys593 at position-3, thereby inhibiting RNA elongation.

O2-4

Analysis of interaction energies for SMAD4 variants bycomputational science
Kazuma Harada 1

Ai Itagaki 1

2133310125b@kindai.ac.jp

1833310128a@kindai.ac.jp

Norihito Kawashita 2
nkawashita@emat.kindai.ac.jp

12

Graduate School of Science and Engineering, Kindai University, 3 -4-1 Kowakae,
Higashiosaka-shi, Osaka, 577-8502, Japan
Faculty of Science and Engineering, Kindai University, 3 -4-1 Kowakae, Higashiosaka-shi
Osaka, 577-8502, Japan

Keywords: SMAD4, Fragment Molecular Orbital Method, protein -protein interaction, mutation
TGF-β (Transforming Growth Factor β) is involved in numerous biological processes, including
regulation of cell proliferation, differentiation, migration and apoptosis [1]. SMAD4 (SmallMothers Against Decapentaplegic 4) mediates this TGF -β signaling, and genetic mutations inSMAD4 have been associated with many cancers, including pancreatic cancer [2]. We havepreviously performed interaction analysis of SMAD4 mutan ts using the fragment molecular orbital(FMO) method [3-4]. In this research, we analyzed the mutations reported subsequently with theaddition of pair interaction decomposition analysis (PIEDA) [5].
The crystal structure used in the calculations was obtained from the Protein Data Bank (PDB) asa wild-type homotrimer (PDBID: 1dd1) [6]. Mutations reported to be pathogenic and likely
pathogenic in ClinVar were introduced the wild-type structure, followed by structure preparationand optimization by MOE. FMO calculations were performed on these structures and compared
before and after mutation introduction using IFIE and PIEDA. ABINIT-MP was used for FMO
calculations, and FMO2-MP2/6-31G* was used as the calculation level.
As a result of comparing the interaction energies for the Arg361 mutants, All five mutations
reported in ClinVar were destabilized by loss of hydrogen bonding w ith Asp537 on adjacent strands,suggesting that they had a profound effect on trimerization. Structural comparison of mutations
within the monomer revealed that Asp351His is destabilized by the loss of hydrogen bond withArg361 and SH-O interaction with Cys363, which may affect the stability of the monomer.
This research was conducted within the activities of the FMO drug design consortium (FMODD);
RIKEN R-CCS Fugaku was used for the FMO calculations (hp220143).
[1] Nakao, A.; et al., TGF-β receptor-mediated signalling through Smad2, Smad3 and Smad4, The
EMBO Journal, 1997, 16(17), 5353-5362.
[2] Ming, Z.; et al., The role of TGF-β/SMAD4 signaling in cancer, Int. J. Biol.Sci., 2018, 14(2),111-123.
[3] Itagaki A.; et al., The 64th Annual Meeting of the Japan Society of Human Genetics, Presentation
No. O3-3.
[4] Kitaura, K.; et al., Pair interaction molecular orbital method: an approximate computational
method for molecular interactions, Chem Phy Lett., 1999, 312, 319-324.
[5] Tsukamoto, T.; et al., Implementation of Pair Interaction Energy Decomposition Analysis and Its
Applications to Protein-Ligand System, J. Conput. Chem.Jpn., 2015, 14(1), 1-9.
[6] Bin, Q.; et al., Crystal structure of a transcriptionally active Smad4 fragment, Structure., 1999,
7,1493-1503.

O2-5Interaction analysis of 14-3-3 and p53
using fragment molecular orbital method
Tatsunori Miyajima 1

Midori Takimoto-Kamimura 22133310123b@kindai.ac.jp

kamimura@cbi-society.org

Norihito Kawashita 3
nkawashita@emat.kindai.ac.jp

123Graduate School of Science and Engineering, Kindai University, 3 -4-1 Kowakae,Higashiosaka-shi, Osaka,577-8502, Japan
Quantum-Structural Life Science Laboratories, CBI Research Institute, Kyowa Create Daiichi build 3F, 3-11-1 Shibaura, Minato, Tokyo, 1008-0023 Japan
Faculty of Science and Engineering, Kindai University, 3-4-1 Kowakae, Higashiosaka-shi,Osaka, 577-8502, Japan

Keywords: 14-3-3, p53, Fragment Molecular Orbital Method
14-3-3σ is associated with several types of cancer and down -regulates malignancy.[1]143-3σ is a positive regulator of p53, one of the tumor suppressor proteins, and protects p53
from proteosome degradation by MDM2 and increases transcriptional activity of p53. [2-3]In this study, we performed interaction analysis of 14-3-3 and p53 complexes with stabilizing
ligand by fragment molecular orbital (FMO) method to provide important information for the designof PPI stabilizers. 14-3-3σ is associated with several types of cancer and down-regulates malignancy.
The 22 crystal structures of structure of 14-3-3 and p53 complex with stabilizing ligand in theliterature [4] were downloaded from Protein Data Bank (PDB). Structural preparation andoptimizations were performed using MOE. We also prepared the complexes that was not registeredin the PDB by MOE Dock. FMO calculations were performed for the structures and the inter fragment interaction energies (IFIE) and pair interaction decomposition analysis (PIEDA) between
each residue and inhibitor were calculated. ABINIT-MP was used for the FMO calculation, andMP2/6-31G* was used as the calculation level.
FMO calculations indicated that the amidine group, which is a common structure of ligand, hasa large electrostatic interaction energy with residues in the 14-3-3 pocket. Structural analysis
confirmed that the amidine group strongly interact with each other by forming salt bridges andhydrogen bonds. PIEDA also suggest that the structures with calculated the dispersion energies withCYS38 and the electrostatic interaction energies with ASP215 in 14-3-3 strongly interact withligand. Therefore, it is considered that designing ligands to have these interactions will make themmore stable stabilizers. In the interaction between 14-3-3 and p53, it was confirmed that the energy
was significantly different by the conformational changes of charged amino acids such as LYS382and ASP391 in p53.
This research was done in activities of the FMO drug d esign consortium (FMODD). For FMO
calculations, RIKEN R-CCS Fugaku were used (hp210130, 220143 quota).
[1] Mhawech, P.; 14-3-3 proteins—an update, Cell Res, 2005, 15, 228–236.
[2] Yang, H.-Y.; et al., 14-3-3 Positively Regulates p53 and Suppresses Tumor Growth, Mol. Cell.
Biol., 2003, 23, 12.
[3] Rayburn, E.; et al., Recent Advances in Validating MDM2 as a Cancer Target, ACAMC, 2009, 9,882–903.
[4] Guillory, X.; et al., Fragment-based Differential Targeting of PPI Stabilizer Interfaces , J. Med.
Chem., 2020, 63, 6694–6707.

O2-6

Comparison of conformer models in PDB structureusing Fragment Molecular Orbital Method

12Yuuki Okayama 1

Norihito Kawashita 1.2

2133310124j@kindai.ac.jp

nkawashita@emat.kindai.ac.jp

Graduate School of Science and Engineering, Kindai University, 3-4-1 Kowakae,
Higashiosaka-shi, Osaka,577-8502, Japan
Faculty of Science and Engineering, Kindai University, 3-4-1 Kowakae, Higashiosaka-shi,Osaka, 577-8502, Japan

Keywords: Protein Data Bank, Conformer, Fragment Molecular Orbital Method, Protein modeling
We are currently performing Fragment Molecular Orbital (FMO) Method [1] calculations on highresolution crystal structure data registered in the PDB, and the problem of conformer model
selection has arisen. When selecting conformers, modeling software usually reads the one with thehighest occupancy as conformer A. However, we found that some structures registered in the PDB
destabilize the energy of the calculation results when Conformer A is selected. Therefore, weperformed FMO calculations on several high-resolution crystal structure data to identify differences
in molecular stability between conformer combinations.
In this study, the high-resolution structure (PDB ID: 1W0N) [2] of the calcium-dependent
carbohydrate-binding module (CBM36) was used to analyze the difference in molecular energy due
to conformational selection.
The crystal structure of CBM36 was downloaded f rom Protein Data Bank (PDB), and four
conformer combinations of Ile12 and Glu128 (A-A, A-B, B-A, and B-B) were prepared. Structuralmodifications and optimizations were performed using MOE2020 (CCG Inc.). FMO calculations
were performed for the created structures and the inter-fragment interaction energies (IFIE) and theenergy components by pair interaction energy decomposition analysis (PIEDA) [3] between Ile12and Glu128 were calculated. ABINIT-MP was used as the FMO calculation, and FMO2-MP2/6-31G
was used as the calculation level.
As a results of the FMO calculations, the interaction energies are very unstable in the structure
between conformer A. This is because the closest atomic distance between conformer A is 2.0 Å.
Comparing the Ile12-Glu128 interaction energies in the other three models showed that only
conformer A-B are stabilized. Thus, it was found that the appropriate selection of conformers should
be made when performing the calculations.
This research was done in activities of the FMO drug design consortium (FMODD). For FMO
calculations, RIKEN R-CCS Fugaku was used (hp220143 quota).
[1] K. Kitaura, et al, Fragment molecular orbital method: use of approximate electrostatic potential ,Chem. Phys. Lett., 2000, 318(6), 614-618.
[2] Sheelan J. T., et al, Ab Initio Structure Determination and Functional Characterization ofCBM36: A New Family of Calcium dependent Carbohydrate Binding Modules., Structure, 2004,12(7), 1177-1187.
[3] K. Kitaura, et al, Pair interaction energy decomposition analysis , J. Comput. Chem., 2007, 28,475-480.