scholarly journals Crystal structure of human METTL6, the m3C methyltransferase

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Ran Chen ◽  
Jie Zhou ◽  
Ling Liu ◽  
Xue-Ling Mao ◽  
Xiaolong Zhou ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Catalytic Mechanism ◽  
Rational Drug Design ◽  
Recognition Mechanism ◽  
Docking Model ◽  
Rational Drug ◽  
Base Ring ◽  
Base Mechanism ◽  
Quaternary Complex ◽  
Key Residues

AbstractIn tRNA, the epigenetic m3C modification at position 32 in the anticodon loop is highly conserved in eukaryotes, which maintains the folding and basepairing functions of the anticodon. However, the responsible enzymes METTL2 and METTL6 were identified only in recent years. The loss of human METTL6 (hMETTL6) affects the translational process and proteostasis in cells, while in mESCs cells, it leads to defective pluripotency potential. Despite its important functions, the catalytic mechanism of the C32 methylation by this enzyme is poorly understood. Here we present the 1.9 Å high-resolution crystal structure of hMETTL6 bound by SAH. The key residues interacting with the ligand were identified and their roles were confirmed by ITC. We generated a docking model for the hMETTL6-SAH-CMP ternary complex. Interestingly, the CMP molecule binds into a cavity in a positive patch with the base ring pointing to the inside, suggesting a flipped-base mechanism for methylation. We further generated a model for the quaternary complex with tRNASer as a component, which reasonably explained the biochemical behaviors of hMETTL6. Taken together, our crystallographic and biochemical studies provide important insight into the molecular recognition mechanism by METTL6 and may aid in the METTL-based rational drug design in the future.

scholarly journals Crystal structure of the N-terminal domain of the trypanosome flagellar protein BILBO1 reveals a ubiquitin fold with a long structured loop for protein binding

2019 ◽  
Vol 295 (6) ◽  
pp. 1489-1499
Author(s):  
Keni Vidilaseris ◽  
Nicolas Landrein ◽  
Yulia Pivovarova ◽  
Johannes Lesigang ◽  
Niran Aeksiri ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Rational Drug Design ◽  
Sub Saharan Africa ◽  
Flagellar Pocket ◽  
Terminal Domain ◽  
Rational Drug ◽  
Cytoplasmic Face ◽  
Flagellar Protein ◽  
Sub Saharan ◽  
Single Flagellum

Trypanosoma brucei is a protist parasite causing sleeping sickness and nagana in sub-Saharan Africa. T. brucei has a single flagellum whose base contains a bulblike invagination of the plasma membrane called the flagellar pocket (FP). Around the neck of the FP on its cytoplasmic face is a structure called the flagellar pocket collar (FPC), which is essential for FP biogenesis. BILBO1 was the first characterized component of the FPC in trypanosomes. BILBO1's N-terminal domain (NTD) plays an essential role in T. brucei FPC biogenesis and is thus vital for the parasite's survival. Here, we report a 1.6-Å resolution crystal structure of TbBILBO1-NTD, which revealed a conserved horseshoe-like hydrophobic pocket formed by an unusually long loop. Results from mutagenesis experiments suggested that another FPC protein, FPC4, interacts with TbBILBO1 by mainly contacting its three conserved aromatic residues Trp-71, Tyr-87, and Phe-89 at the center of this pocket. Our findings disclose the binding site of TbFPC4 on TbBILBO1-NTD, which may provide a basis for rational drug design targeting BILBO1 to combat T. brucei infections.

scholarly journals Atomic-Level Investigation of Reactant Recognition Mechanism and Thermodynamic Property in Glucosamine 6-Phosphate Deaminase Catalysis

2021 ◽  
Vol 9 ◽  
Author(s):  
Xiao Zhang ◽  
Xiaoyuan Liu ◽  
Zhiyang Zhang ◽  
Yuan Zhao ◽  
Chaojie Wang
Keyword(s):  
Hydrogen Bond ◽  
Active Site ◽  
Catalytic Mechanism ◽  
Polar Solvent ◽  
Enzymatic Reaction ◽  
Alkaline Condition ◽  
Recognition Mechanism ◽  
Hydrogen Bond Interactions ◽  
Dynamics Simulations ◽  
Key Residues

Glucosamine 6-phosphate deaminase (NagB) influences the direction of N-acetylglucosamine (GlcNAc) metabolism, facilitating the conversion of D-glucosamine 6-phosphate (GlcN6P) to D-fructose 6-phosphate (Fru6P) with the release of ammonia. Here, extensive molecular dynamics simulations combined with various techniques were performed to study the recognition and delivery process of GlcN6P by SmuNagB, due to its guidance of subsequent enzymatic reaction. The key residues Lys194, His130, Arg127, Thr38, and Ser37 stabilize GlcN6P in the active site by hydrogen bond interactions, therein electrostatic and polar solvent effects provide the primary traction. Four delivery channels were identified, with GlcN6P most likely to enter the active site of NagB through a “door” comprising residues 6–10, 122–136, and 222–233. The corresponding mechanism and thermodynamic properties were investigated. An exothermic recognition and delivery process were detected, accompanied by the flipping of GlcN6P and changes in key direct and indirect hydrogen bond interactions, which provide the driving force for the chemical reaction to occur. Furthermore, “the lid motif” was identified that remain open in alkaline condition with different extent of opening at each stage of transfer that induced GlcN6P to move the active site of NagB. The work will assist in the elucidation of the catalytic mechanism of action of NagB, allowing inhibitors to be designed with superior dynamic behavior.

scholarly journals The Crystal Structure of the Spodoptera litura Chemosensory Protein CSP8

Insects ◽  
2021 ◽  
Vol 12 (7) ◽  
pp. 602
Author(s):  
Qian Jia ◽  
Hui Zeng ◽  
Jinbing Zhang ◽  
Shangfang Gao ◽  
Nan Xiao ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Spodoptera Litura ◽  
Spherical Shape ◽  
Rational Drug Design ◽  
Structural Basis ◽  
Binding Assays ◽  
Recognition Mechanism ◽  
Generalist Herbivore ◽  
Chemosensory Protein ◽  
Critical Components

Spodoptera litura F. is a generalist herbivore and one of the most important economic pests feeding on about 300 host plants in many Asian countries. Specific insect behaviors can be stimulated after recognizing chemicals in the external environment through conserved chemosensory proteins (CSPs) in chemoreceptive organs, which are critical components of the olfactory systems. To explore its structural basis for ligand-recognizing capability, we solved the 2.3 Å crystal structure of the apoprotein of S. litura CSP8 (SlCSP8). The SlCSP8 protein displays a conserved spherical shape with a negatively charged surface. Our binding assays showed that SlCSP8 bound several candidate ligands with differential affinities, with rhodojaponin III being the most tightly bound ligand. Our crystallographic and biochemical studies provide important insight into the molecular recognition mechanism of the sensory protein SlCSP8 and the CSP family in general, and they suggest that CSP8 is critical for insects to identify rhodojaponin III, which may aid in the CSP-based rational drug design in the future.

scholarly journals Crystal structure of the human PRPK–TPRKB complex

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Jian Li ◽  
Xinli Ma ◽  
Surajit Banerjee ◽  
Hanyong Chen ◽  
Weiya Ma ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Structural Model ◽  
Rational Drug Design ◽  
Activation Loop ◽  
Active Conformation ◽  
Kinase Activation ◽  
Disease Mutations ◽  
Rational Drug ◽  
Anti Cancer ◽  
Keops Complex

AbstractMutations of the p53-related protein kinase (PRPK) and TP53RK-binding protein (TPRKB) cause Galloway-Mowat syndrome (GAMOS) and are found in various human cancers. We have previously shown that small compounds targeting PRPK showed anti-cancer activity against colon and skin cancer. Here we present the 2.53 Å crystal structure of the human PRPK-TPRKB-AMPPNP (adenylyl-imidodiphosphate) complex. The structure reveals details in PRPK-AMPPNP coordination and PRPK-TPRKB interaction. PRPK appears in an active conformation, albeit lacking the conventional kinase activation loop. We constructed a structural model of the human EKC/KEOPS complex, composed of PRPK, TPRKB, OSGEP, LAGE3, and GON7. Disease mutations in PRPK and TPRKB are mapped into the structure, and we show that one mutation, PRPK K238Nfs*2, lost the binding to OSGEP. Our structure also makes the virtual screening possible and paves the way for more rational drug design.

scholarly journals A structural study of TatD from Staphylococcus aureus elucidates a putative DNA-binding mode of a Mg2+-dependent nuclease

IUCrJ ◽  
2020 ◽  
Vol 7 (3) ◽  
pp. 509-521
Author(s):  
Kyu-Yeon Lee ◽  
Seung-Ho Cheon ◽  
Dong-Gyun Kim ◽  
Sang Jae Lee ◽  
Bong-Jin Lee
Keyword(s):  
Crystal Structure ◽  
Staphylococcus Aureus ◽  
Dna Binding ◽  
Catalytic Mechanism ◽  
Nuclease Activity ◽  
Binding Mode ◽  
Functional Study ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Key Residues

TatD has been thoroughly investigated as a DNA-repair enzyme and an apoptotic nuclease, and still-unknown TatD-related DNases are considered to play crucial cellular roles. However, studies of TatD from Gram-positive bacteria have been hindered by an absence of atomic detail and the resulting inability to determine function from structure. In this study, an X-ray crystal structure of SAV0491, which is the TatD enzyme from the Gram-positive bacterium Staphylococcus aureus (SaTatD), is reported at a high resolution of 1.85 Å with a detailed atomic description. Although SaTatD has the common TIM-barrel fold shared by most TatD-related homologs, and PDB entry 2gzx shares 100% sequence identity with SAV0491, the crystal structure of SaTatD revealed a unique binding mode of two phosphates interacting with two Ni2+ ions. Through a functional study, it was verified that SaTatD has Mg2+-dependent nuclease activity as a DNase and an RNase. In addition, structural comparison with TatD homologs and the identification of key residues contributing to the binding mode of Ni2+ ions and phosphates allowed mutational studies to be performed that revealed the catalytic mechanism of SaTatD. Among the key residues composing the active site, the acidic residues Glu92 and Glu202 had a critical impact on catalysis by SaTatD. Furthermore, based on the binding mode of the two phosphates and structural insights, a putative DNA-binding mode of SaTatD was proposed using in silico docking. Overall, these findings may serve as a good basis for understanding the relationship between the structure and function of TatD proteins from Gram-positive bacteria and may provide critical insights into the DNA-binding mode of SaTatD.

scholarly journals Computational Determination of Potential Multiprotein Targeting Natural Compounds for Rational Drug Design Against SARS-COV-2

Molecules ◽  
2021 ◽  
Vol 26 (3) ◽  
pp. 674
Author(s):  
Ziyad Tariq Muhseen ◽  
Alaa R. Hameed ◽  
Halah M. H. Al-Hasani ◽  
Sajjad Ahmad ◽  
Guanglin Li
Keyword(s):  
Free Energy ◽  
Binding Energy ◽  
Binding Free Energy ◽  
Binding Mode ◽  
Natural Compounds ◽  
Rational Drug Design ◽  
Dynamics Simulation ◽  
Antiviral Compounds ◽  
Rational Drug ◽  
Key Residues

SARS-CoV-2 caused the current COVID-19 pandemic and there is an urgent need to explore effective therapeutics that can inhibit enzymes that are imperative in virus reproduction. To this end, we computationally investigated the MPD3 phytochemical database along with the pool of reported natural antiviral compounds with potential to be used as anti-SARS-CoV-2. The docking results demonstrated glycyrrhizin followed by azadirachtanin, mycophenolic acid, kushenol-w and 6-azauridine, as potential candidates. Glycyrrhizin depicted very stable binding mode to the active pocket of the Mpro (binding energy, −8.7 kcal/mol), PLpro (binding energy, −7.9 kcal/mol), and Nucleocapsid (binding energy, −7.9 kcal/mol) enzymes. This compound showed binding with several key residues that are critical to natural substrate binding and functionality to all the receptors. To test docking prediction, the compound with each receptor was subjected to molecular dynamics simulation to characterize the molecule stability and decipher its possible mechanism of binding. Each complex concludes that the receptor dynamics are stable (Mpro (mean RMSD, 0.93 Å), PLpro (mean RMSD, 0.96 Å), and Nucleocapsid (mean RMSD, 3.48 Å)). Moreover, binding free energy analyses such as MMGB/PBSA and WaterSwap were run over selected trajectory snapshots to affirm intermolecular affinity in the complexes. Glycyrrhizin was rescored to form strong affinity complexes with the virus enzymes: Mpro (MMGBSA, −24.42 kcal/mol and MMPBSA, −10.80 kcal/mol), PLpro (MMGBSA, −48.69 kcal/mol and MMPBSA, −38.17 kcal/mol) and Nucleocapsid (MMGBSA, −30.05 kcal/mol and MMPBSA, −25.95 kcal/mol), were dominated mainly by vigorous van der Waals energy. Further affirmation was achieved by WaterSwap absolute binding free energy that concluded all the complexes in good equilibrium and stability (Mpro (mean, −22.44 kcal/mol), PLpro (mean, −25.46 kcal/mol), and Nucleocapsid (mean, −23.30 kcal/mol)). These promising findings substantially advance our understanding of how natural compounds could be shaped to counter SARS-CoV-2 infection.

Structural and biochemical analyses of theStreptococcus pneumoniaeL,D-carboxypeptidase DacB

2015 ◽  
Vol 71 (2) ◽  
pp. 283-292
Author(s):  
Juan Zhang ◽  
Yi-Hu Yang ◽  
Yong-Liang Jiang ◽  
Cong-Zhao Zhou ◽  
Yuxing Chen
Keyword(s):  
Crystal Structure ◽  
Molecular Docking ◽  
Catalytic Mechanism ◽  
Substrate Recognition ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Binding Properties ◽  
Biochemical Analyses ◽  
Key Residues ◽  
Structural Insights

The L,D-carboxypeptidase DacB plays a key role in the remodelling ofStreptococcus pneumoniaepeptidoglycan during cell division. In order to decipher its substrate-binding properties and catalytic mechanism, the 1.71 Å resolution crystal structure of DacB fromS. pneumoniaeTIGR4 is reported. Structural analyses in combination with comparisons with the recently reported structures of DacB fromS. pneumoniaeD39 and R6 clearly demonstrate that DacB adopts a zinc-dependent carboxypeptidase fold and belongs to the metallopeptidase M15B subfamily. In addition, enzymatic activity assays further confirm that DacB indeed acts as an L,D-carboxypeptidase towards the tetrapeptide L-Ala-D-iGln-L-Lys-D-Ala of the peptidoglycan stem, withKmandkcatvalues of 2.84 ± 0.37 mMand 91.49 ± 0.05 s−1, respectively. Subsequent molecular docking and site-directed mutagenesis enable the assignment of the key residues that bind to the tetrapeptide. Altogether, these findings provide structural insights into substrate recognition in the metallopeptidase M15B subfamily.

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