Modification of the active site of Mycobacterium tuberculosis KatG after disruption of the Met–Tyr–Trp cross-linked adduct

Kapetanaki, Sofia M.; Zhao, Xiangbo; Yu, Shengwei; Magliozzo, Richard S.; Schelvis, Johannes P. M.

doi:10.1016/j.jinorgbio.2006.11.004

Cited by 11 publications

(7 citation statements)

References 78 publications

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“…ARG104, TRP107, and HIS108 were located in the distal pocket of the active site. There are covalent linkages between TRP107 and TYR229 [23,24] . As shown in Figure 3, compound 1 , compound 2 , isoniazid, and pyrazinamide can all be accommodated in the binding pocket of Mtb KatG.…”

Section: Resultsmentioning

confidence: 99%

Antimycobacterial Compound of Cynoglossum lanceolatum Forsk.: Bioassay Guided Isolation, Molecular Docking, Synthesis of Analogs, and a Plausible Mechanism of Action

Yang

Xin

Wang

et al. 2023

Chemistry & Biodiversity

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Tuberculosis (TB) remains a major threat to human health. Due to the prevalence of drug‐resistant Mycobacterium tuberculosis (Mtb), it is urgent to discover drugs with new mechanisms of action (MOA) to ensure effectiveness against strains that are resistant to existing TB drugs. Cynoglossum lanceolatum Forsk was used to treat TB in Traditional Chinese Medicine. In this article, shikonin, the anti‐Mtb active component, was obtained from the whole herb extract of C. lanceolatum by bioassay‐guided isolation. Using the microplate alamar blue assay (MABA), the minimum inhibitory concentration (MIC) of shikonin against Mtb was determined to be 128 μg/mL. In order to obtain a more efficient anti‐Mtb molecule, (E)‐1‐(6‐bromo‐2,3‐dihydrochromen‐4‐ylidene)thiosemicarbazide was synthesized based on the scaffold of shikonin, which exhibited potent activity against Mtb (MIC=4 μg/mL). These results highlight that both naphthalene‐1,4‐dione and chroman‐4‐one are pharmacophores with activities against Mtb. To investigate a plausible mechanism of action, the molecular docking was firstly performed against catalase‐peroxidase enzyme (KatG) of Mtb using AutoDock 4 software. The results demonstrated that both shikonin and (E)‐1‐(6‐bromo‐2,3‐dihydrochromen‐4‐ylidene)thiosemicarbazide could bind to the active site of Mtb KatG. KatG enzyme activity and intracellular reactive oxygen species (ROS) levels in Mtb cells were then measured by ultraviolet spectrophotometric method and fluorescence microplate reader assay, respectively. The experiments confirmed that above compounds could inhibit the catalytic activity of Mtb KatG, and cause the ROS accumulation in Mtb cells. Therefore, inhibition of KatG may be a novel mechanism of action for these two compounds to fight against Mtb.

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Section: Resultsmentioning

confidence: 99%

Antimycobacterial Compound of Cynoglossum lanceolatum Forsk.: Bioassay Guided Isolation, Molecular Docking, Synthesis of Analogs, and a Plausible Mechanism of Action

Yang

Xin

Wang

et al. 2023

Chemistry & Biodiversity

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“…The X-ray structures revealed that KatGs possess unique covalent bonds formed among the side chains of three distal residues, Met-Tyr-Trp, which are located on the distal side of the haem active site. Mutagenesis studies confirmed that the Met-Tyr-Trp cross-link is required for catalatic activity (Regelsberger et al, 2000;Jakopitsch et al, 2004;Ghiladi et al, 2005;Kapetanaki et al, 2007;Zhao et al, 2010Zhao et al, , 2013. These structures have provided many insights into the structure and function of KatGs.…”

Section: Introductionmentioning

confidence: 82%

The 2.2 Å resolution structure of the catalase-peroxidase KatG fromSynechococcus elongatusPCC7942

Kamachi¹,

Wada²,

Tamoi³

et al. 2014

Acta Cryst Sect F

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The crystal structure of catalase-peroxidase from Synechococcus elongatus PCC7942 (SeKatG) was solved by molecular replacement and refined to an Rwork of 16.8% and an Rfree of 20.6% at 2.2 Å resolution. The asymmetric unit consisted of only one subunit of the catalase-peroxidase molecule, including a protoporphyrin IX haem moiety and two sodium ions. A typical KatG covalent adduct was formed, Met248-Tyr222-Trp94, which is a key structural element for catalase activity. The crystallographic equivalent subunit was created by a twofold symmetry operation to form the functional dimer. The overall structure of the dimer was quite similar to other KatGs. One sodium ion was located close to the proximal Trp314. The location and configuration of the proximal cation site were very similar to those of typical peroxidases such as ascorbate peroxidase. These features may provide a structural basis for the behaviour of the radical localization/delocalization during the course of the enzymatic reaction.

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“…The Ser315Thr substitution in KatG impacts on the shifting of substrate binding chan nel from 6.0 to 4.7 Å [27]. Consequently the mutant failed to bind the INH, and subsequently decreased 160 times in the INH activation compared with KatGwt [14].…”

Section: Fig 4 Electrophoregram Of Katg Protein In Sds Page Proteimentioning

confidence: 99%

“…The mutant KatG Ser315Thr, that is commonly found in clinical isolates and associated with INH resistance, decreases the activities of catalaseper oxidase and INH oxidation [13,14]. The amino acid Ser315 in KatG is closely put in the active environ ment.…”

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confidence: 99%

Mutation of katG in a clinical isolate of Mycobacterium tuberculosis: effects on catalase-peroxidase for isoniazid activation

Purkan¹,

Ihsanawati²,

Natalia³

et al. 2016

Ukr.Biochem.J.

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Mutations in katG gene are often associated with isoniazid (INH) resistance in Mycobacterium tuberculosis strain. This research was perfomed to identify the katG mutation in clinical isolate (L8) that is resistant to INH at 1 μg/ml. In addition to characterize the catalase-peroxidase of KatG L8 and perform the ab initio structural study of the protein to get a more complete understanding in drug activation and the resistance mechanism. The katG gene was cloned and expressed in Escherichia coli, then followed by characterization of catalase-peroxidase of KatG. The structure modelling was performed to know a basis of alterations in enzyme activity. A substitution of A713G that correspond to Asn238Ser replacement was found in the L8 katG. The Asn238Ser modification leads to a decline in the activity of catalase-peroxidase and INH oxidation of the L8 KatG protein. The catalytic efficiency (Kcat/KM) of mutant KatGAsn238Ser respectively decreases to 41 and 52% for catalase and peroxidase. The mutant KatGAsn238Ser also shows a decrease of 62% in INH oxidation if compared to a wild type KatG (KatGwt). The mutant Asn238Ser might cause instability in the substrate binding site of KatG, because of removal of a salt bridge connecting the amine group of Asn238 to the carboxyl group of Glu233, which presents in KatGwt. The lost of the salt bridge in the substrate binding site in mutant KatGAsn238Ser created changes unfavorable for enzyme activities, which in turn emerge as INH resistance in the L8 isolate of M. tuberculosis.

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Modification of the active site of Mycobacterium tuberculosis KatG after disruption of the Met–Tyr–Trp cross-linked adduct

Cited by 11 publications

References 78 publications

Antimycobacterial Compound of Cynoglossum lanceolatum Forsk.: Bioassay Guided Isolation, Molecular Docking, Synthesis of Analogs, and a Plausible Mechanism of Action

Antimycobacterial Compound of Cynoglossum lanceolatum Forsk.: Bioassay Guided Isolation, Molecular Docking, Synthesis of Analogs, and a Plausible Mechanism of Action

The 2.2 Å resolution structure of the catalase-peroxidase KatG fromSynechococcus elongatusPCC7942

Mutation of katG in a clinical isolate of Mycobacterium tuberculosis: effects on catalase-peroxidase for isoniazid activation

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