scholarly journals Structural analysis of mycobacterial homoserine transacetylases central to methionine biosynthesis reveals druggable active site

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Catherine T. Chaton ◽  
Emily S. Rodriguez ◽  
Robert W. Reed ◽  
Jian Li ◽  
Cameron W. Kenner ◽  
...  
Keyword(s):  
Infectious Disease ◽  
Mycobacterium Tuberculosis ◽  
Active Site ◽  
Fungal Species ◽  
Biosynthesis Pathway ◽  
Methionine Biosynthesis ◽  
Structure Based Drug Design ◽  
High Degree ◽  
Homoserine Transacetylase ◽  
Host Metabolism

AbstractMycobacterium tuberculosis is the cause of the world’s most deadly infectious disease. Efforts are underway to target the methionine biosynthesis pathway, as it is not part of the host metabolism. The homoserine transacetylase MetX converts l-homoserine to O-acetyl-l-homoserine at the committed step of this pathway. In order to facilitate structure-based drug design, we determined the high-resolution crystal structures of three MetX proteins, including M. tuberculosis (MtMetX), Mycolicibacterium abscessus (MaMetX), and Mycolicibacterium hassiacum (MhMetX). A comparison of homoserine transacetylases from other bacterial and fungal species reveals a high degree of structural conservation amongst the enzymes. Utilizing homologous structures with bound cofactors, we analyzed the potential ligandability of MetX. The deep active-site tunnel surrounding the catalytic serine yielded many consensus clusters during mapping, suggesting that MtMetX is highly druggable.

scholarly journals Structural analysis of mycobacterial homoserine transacetylases central to methionine biosynthesis reveals druggable active site

2019 ◽  
Author(s):  
Catherine T. Chaton ◽  
Emily S. Rodriguez ◽  
Robert W. Reed ◽  
Jian Li ◽  
Cameron W. Kenner ◽  
...  
Keyword(s):  
Infectious Disease ◽  
Mycobacterium Tuberculosis ◽  
Active Site ◽  
Fungal Species ◽  
Biosynthesis Pathway ◽  
Methionine Biosynthesis ◽  
Structure Based Drug Design ◽  
High Degree ◽  
Homoserine Transacetylase ◽  
Host Metabolism

AbstractMycobacterium tuberculosis is the cause of the world’s most deadly infectious disease. Efforts are underway to target the methionine biosynthesis pathway, as it is not part of the host metabolism. The homoserine transacetylase MetX converts L-homoserine to O-acetyl-L-homoserine at the committed step of this pathway. In order to facilitate structure-based drug design, we determined the high-resolution crystal structures of three MetX proteins, including M. tuberculosis (MtMetX), Mycolicibacterium abscessus (MaMetX), and Mycolicibacterium hassiacum (MhMetX). A comparison of homoserine transacetylases from other bacterial and fungal species reveals a high degree of structural conservation amongst the enzymes. Utilizing homologous structures with bound cofactors, we analyzed the potential ligandability of MetX. The deep active-site tunnel surrounding the catalytic serine yielded many consensus clusters during mapping, suggesting that MtMetX is highly druggable.

scholarly journals Allosteric regulation of menaquinone (vitamin K2) biosynthesis in the human pathogen Mycobacterium tuberculosis

2020 ◽  
Vol 295 (12) ◽  
pp. 3759-3770 ◽  
Author(s):  
Ghader Bashiri ◽  
Laura V. Nigon ◽  
Ehab N. M. Jirgis ◽  
Ngoc Anh Thu Ho ◽  
Tamsyn Stanborough ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Active Site ◽  
Feedback Inhibition ◽  
Feedback Regulation ◽  
Signal Propagation ◽  
Biosynthesis Pathway ◽  
Human Pathogen ◽  
Energy Status ◽  
Allosteric Site ◽  
Arginine Residues

Menaquinone (vitamin K2) plays a vital role in energy generation and environmental adaptation in many bacteria, including the human pathogen Mycobacterium tuberculosis (Mtb). Although menaquinone levels are known to be tightly linked to the cellular redox/energy status of the cell, the regulatory mechanisms underpinning this phenomenon are unclear. The first committed step in menaquinone biosynthesis is catalyzed by MenD, a thiamine diphosphate–dependent enzyme comprising three domains. Domains I and III form the MenD active site, but no function has yet been ascribed to domain II. Here, we show that the last cytosolic metabolite in the menaquinone biosynthesis pathway, 1,4-dihydroxy-2-naphthoic acid (DHNA), binds to domain II of Mtb-MenD and inhibits its activity. Using X-ray crystallography of four apo- and cofactor-bound Mtb-MenD structures, along with several spectroscopy assays, we identified three arginine residues (Arg-97, Arg-277, and Arg-303) that are important for both enzyme activity and the feedback inhibition by DHNA. Among these residues, Arg-277 appeared to be particularly important for signal propagation from the allosteric site to the active site. This is the first evidence of feedback regulation of the menaquinone biosynthesis pathway in bacteria, identifying a protein-level regulatory mechanism that controls menaquinone levels within the cell and may therefore represent a good target for disrupting menaquinone biosynthesis in M. tuberculosis.

scholarly journals Chemical validation of Mycobacterium tuberculosis phosphopantetheine adenylyltransferase using fragment linking and CRISPR interference

2020 ◽  
Author(s):  
Jamal El Bakali ◽  
Michal Blaszczyk ◽  
Joanna C. Evans ◽  
Jennifer A. Boland ◽  
William J. McCarthy ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Crystal Structures ◽  
Active Site ◽  
Drug Target ◽  
Potential Drug Target ◽  
Biosynthesis Pathway ◽  
Potential Drug ◽  
Antimicrobial Drugs ◽  
X Ray ◽  
Crispr Interference

AbstractThe coenzyme A (CoA) biosynthesis pathway has attracted attention as a potential target for much-needed novel antimicrobial drugs, including for the treatment of tuberculosis (TB), the lethal disease caused by Mycobacterium tuberculosis (Mtb). Seeking to identify the first inhibitors of Mtb phosphopantetheine adenylyltransferase (MtbPPAT), the enzyme that catalyses the penultimate step in CoA biosynthesis, we performed a fragment screen. In doing so, we discovered three series of fragments that occupy distinct regions of the MtbPPAT active site, presenting a unique opportunity for fragment linking. Here we show how, guided by X-ray crystal structures, we could link weakly-binding fragments to produce an active site binder with a KD < 20 μM and on-target anti-Mtb activity, as demonstrated using CRISPR interference. This study represents a big step toward validating MtbPPAT as a potential drug target and designing a MtbPPAT-targeting anti-TB drug.Abstract Figure

scholarly journals DOCKING STUDIES FOR VARIOUS ANTIBACTERIAL BENZILATE DERIVATIVES

2017 ◽  
Vol 10 (4) ◽  
pp. 268
Author(s):  
Sudha Rajendran ◽  
Brindha Devi P ◽  
Charles C Kanakam ◽  
Nithya G
Keyword(s):  
Infectious Disease ◽  
Amino Acids ◽  
Mycobacterium Tuberculosis ◽  
Active Site ◽  
Docking Studies ◽  
Antibacterial Drug ◽  
Structure Activity ◽  
Docking Tool ◽  
Excretion Study ◽  
Benzilic Acid

Objectives: In this study, we have focused on discovering the leads for the enzyme targets of infectious disease tuberculosis. We employed computeraided drug design docking tool,to discover new leads for Mycobacterium tuberculosis (MTB).Methods: Five compounds were synthesized and they are made to dock into the active site of the enzyme; retrieved from protein data bank.Results: The docking studies and structure–activity relationship reveals that the compound 2’-chloro-4-methoxy-3nitro benzilic acid after threedifferent docking strategies reveals that the score was found to be higher compared with others(−5.568 kcal/mol).Conclusion: On the closer analysis of this molecule, the molecule showed stacking interaction and the compound has also found to be surrounded by non-polar amino acids, which makes this molecule potent toward antibacterial drug discovery.Keywords: Antibacterials, Docking, Absorption, Distribution, Metabolism and excretion study, Resistance.

Tissue section lifting for electron microscopy

1978 ◽  
Vol 36 (2) ◽  
pp. 86-87
Author(s):  
Adrian F. van Dellen
Keyword(s):  
Infectious Disease ◽  
Electron Microscopy ◽  
Tissue Culture ◽  
Organ System ◽  
Surrounding Tissue ◽  
Paraffin Sections ◽  
Surface Areas ◽  
Specific Determination ◽  
High Degree ◽  
Accuracy And Repeatability

The morphologic pathologist may require information on the ultrastructure of a non-specific lesion seen under the light microscope before he can make a specific determination. Such lesions, when caused by infectious disease agents, may be sparsely distributed in any organ system. Tissue culture systems, too, may only have widely dispersed foci suitable for ultrastructural study. In these situations, when only a few, small foci in large tissue areas are useful for electron microscopy, it is advantageous to employ a methodology which rapidly selects a single tissue focus that is expected to yield beneficial ultrastructural data from amongst the surrounding tissue. This is in essence what "LIFTING" accomplishes. We have developed LIFTING to a high degree of accuracy and repeatability utilizing the Microlift (Fig 1), and have successfully applied it to tissue culture monolayers, histologic paraffin sections, and tissue blocks with large surface areas that had been initially fixed for either light or electron microscopy.

scholarly journals Ophiocordyceps salganeicola, a parasite of social cockroaches in Japan and insights into the evolution of other closely-related Blattodea-associated lineages

IMA Fungus ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
João P. M. Araújo ◽  
Mitsuru G. Moriguchi ◽  
Shigeru Uchiyama ◽  
Noriko Kinjo ◽  
Yu Matsuura
Keyword(s):  
Genetic Similarity ◽  
Evolutionary History ◽  
Fungal Species ◽  
Evolutionary Relationships ◽  
Ryukyu Archipelago ◽  
Phylogenetic Placement ◽  
The Social ◽  
Host Jump ◽  
High Degree ◽  
A New Species

AbstractThe entomopathogenic genus Ophiocordyceps includes a highly diverse group of fungal species, predominantly parasitizing insects in the orders Coleoptera, Hemiptera, Hymenoptera and Lepidoptera. However, other insect orders are also parasitized by these fungi, for example the Blattodea (termites and cockroaches). Despite their ubiquity in nearly all environments insects occur, blattodeans are rarely found infected by filamentous fungi and thus, their ecology and evolutionary history remain obscure. In this study, we propose a new species of Ophiocordyceps infecting the social cockroaches Salganea esakii and S. taiwanensis, based on 16 years of collections and field observations in Japan, especially in the Ryukyu Archipelago. We found a high degree of genetic similarity between specimens from different islands, infecting these two Salganea species and that this relationship is ancient, likely not originating from a recent host jump. Furthermore, we found that Ophiocordyceps lineages infecting cockroaches evolved around the same time, at least twice, one from beetles and the other from termites. We have also investigated the evolutionary relationships between Ophiocordyceps and termites and present the phylogenetic placement of O. cf. blattae. Our analyses also show that O. sinensis could have originated from an ancestor infecting termite, instead of beetle larvae as previously proposed.

scholarly journals Morpho-molecular identification and first report of Fusarium equiseti in causing chilli wilt from Kashmir (Northern Himalayas)

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ammarah Hami ◽  
Rovidha S. Rasool ◽  
Nisar A. Khan ◽  
Sheikh Mansoor ◽  
Mudasir A. Mir ◽  
...  
Keyword(s):  
Dna Barcoding ◽  
Pcr Amplification ◽  
Its Region ◽  
Fungal Species ◽  
Pathogenicity Test ◽  
Kashmir Valley ◽  
Fusarium Equiseti ◽  
Infected Root ◽  
Fusarium Sp ◽  
High Degree

AbstractChilli (Capsicum annuum L.) is one of the most significant vegetable and spice crop. Wilt caused by Fusarium Sp. has emerged as a serious problem in chilli production. Internal transcribed spacer (ITS) region is widely used as a DNA barcoding marker to characterize the diversity and composition of Fusarium communities. ITS regions are heavily used in both molecular methods and ecological studies of fungi, because of its high degree of interspecific variability, conserved primer sites and multiple copy nature in the genome. In the present study we focused on morphological and molecular characterization of pathogen causing chilli wilt. Chilli plants were collected from four districts of Kashmir valley of Himalayan region. Pathogens were isolated from infected root and stem of the plants. Isolated pathogens were subjected to DNA extraction and PCR amplification. The amplified product was sequenced and three different wilt causing fungal isolates were obtained which are reported in the current investigation. In addition to Fusarium oxysporum and Fusarium solani, a new fungal species was found in association with the chilli wilt in Kashmir valley viz., Fusarium equiseti that has never been reported before from this region. The studies were confirmed by pathogenicity test and re-confirmation by DNA barcoding.

scholarly journals Toward the Identification of Potential α-Ketoamide Covalent Inhibitors for SARS-CoV-2 Main Protease: Fragment-Based Drug Design and MM-PBSA Calculations

Processes ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 1004
Author(s):  
Mahmoud A. El Hassab ◽  
Mohamed Fares ◽  
Mohammed K. Abdel-Hamid Amin ◽  
Sara T. Al-Rashood ◽  
Amal Alharbi ◽  
...  
Keyword(s):  
Drug Design ◽  
Active Site ◽  
Binding Free Energy ◽  
Stable Complex ◽  
Active Site Residues ◽  
Structure Based Drug Design ◽  
Enzyme Active Site ◽  
Potential Inhibitor ◽  
Main Protease ◽  
Covalent Docking

Since December 2019, the world has been facing the outbreak of the SARS-CoV-2 pandemic that has infected more than 149 million and killed 3.1 million people by 27 April 2021, according to WHO statistics. Safety measures and precautions taken by many countries seem insufficient, especially with no specific approved drugs against the virus. This has created an urgent need to fast track the development of new medication against the virus in order to alleviate the problem and meet public expectations. The SARS-CoV-2 3CL main protease (Mpro) is one of the most attractive targets in the virus life cycle, which is responsible for the processing of the viral polyprotein and is a key for the ribosomal translation of the SARS-CoV-2 genome. In this work, we targeted this enzyme through a structure-based drug design (SBDD) protocol, which aimed at the design of a new potential inhibitor for Mpro. The protocol involves three major steps: fragment-based drug design (FBDD), covalent docking and molecular dynamics (MD) simulation with the calculation of the designed molecule binding free energy at a high level of theory. The FBDD step identified five molecular fragments, which were linked via a suitable carbon linker, to construct our designed compound RMH148. The mode of binding and initial interactions between RMH148 and the enzyme active site was established in the second step of our protocol via covalent docking. The final step involved the use of MD simulations to test for the stability of the docked RMH148 into the Mpro active site and included precise calculations for potential interactions with active site residues and binding free energies. The results introduced RMH148 as a potential inhibitor for the SARS-CoV-2 Mpro enzyme, which was able to achieve various interactions with the enzyme and forms a highly stable complex at the active site even better than the co-crystalized reference.

scholarly journals Threonine 57 is required for the post-translational activation ofEscherichia coliaspartate α-decarboxylase

2014 ◽  
Vol 70 (4) ◽  
pp. 1166-1172 ◽  
Author(s):  
Michael E. Webb ◽  
Briony A. Yorke ◽  
Tom Kershaw ◽  
Sarah Lovelock ◽  
Carina M. C. Lobley ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Active Site ◽  
Site Directed Mutagenesis ◽  
Intramolecular Rearrangement ◽  
Directed Mutagenesis ◽  
Vitamin B ◽  
Biosynthesis Pathway ◽  
Serine Residue ◽  
Translational Activation

Aspartate α-decarboxylase is a pyruvoyl-dependent decarboxylase required for the production of β-alanine in the bacterial pantothenate (vitamin B5) biosynthesis pathway. The pyruvoyl group is formedviathe intramolecular rearrangement of a serine residue to generate a backbone ester intermediate which is cleaved to generate an N-terminal pyruvoyl group. Site-directed mutagenesis of residues adjacent to the active site, including Tyr22, Thr57 and Tyr58, reveals that only mutation of Thr57 leads to changes in the degree of post-translational activation. The crystal structure of the site-directed mutant T57V is consistent with a non-rearranged backbone, supporting the hypothesis that Thr57 is required for the formation of the ester intermediate in activation.

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