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Solution-state NMR structure and biophysical characterization of zinc-substituted rubredoxin B (Rv3250c) from Mycobacterium tuberculosis

Owing to the evolution of multi-drug-resistant and extremely drug-resistant Mycobacterium tuberculosis strains, there is an urgent need to develop new antituberculosis strategies to prevent TB epidemics in the industrial world. Among the potential new drug targets are two small nonheme iron-binding...

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Autores principales: Buchko, Garry W., Hewitt, Stephen N., Napuli, Alberto J., Van Voorhis, Wesley C., Myler, Peter J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: International Union of Crystallography 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3169417/
https://www.ncbi.nlm.nih.gov/pubmed/21904065
http://dx.doi.org/10.1107/S1744309111008189
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author Buchko, Garry W.
Hewitt, Stephen N.
Napuli, Alberto J.
Van Voorhis, Wesley C.
Myler, Peter J.
author_facet Buchko, Garry W.
Hewitt, Stephen N.
Napuli, Alberto J.
Van Voorhis, Wesley C.
Myler, Peter J.
author_sort Buchko, Garry W.
collection PubMed
description Owing to the evolution of multi-drug-resistant and extremely drug-resistant Mycobacterium tuberculosis strains, there is an urgent need to develop new antituberculosis strategies to prevent TB epidemics in the industrial world. Among the potential new drug targets are two small nonheme iron-binding proteins, rubredoxin A (Rv3251c) and rubredoxin B (Rv3250c), which are believed to play a role in electron-transfer processes. Here, the solution structure and biophysical properties of one of these two proteins, rubredoxin B (Mt-RubB), determined in the zinc-substituted form are reported. The zinc-substituted protein was prepared by expressing Mt-RubB in minimal medium containing excess zinc acetate. Size-exclusion chromatography and NMR spectroscopy indicated that Mt-RubB was a monomer in solution. The structure (PDB entry 2kn9) was generally similar to those of other rubredoxins, containing a three-stranded anti­parallel β-sheet (β2–β1–β3) and a metal tetrahedrally coordinated to the S atoms of four cysteine residues (Cys9, Cys12, Cys42 and Cys45). The first pair of cysteine residues is at the C-terminal end of the first β-­strand and the second pair of cysteine residues is towards the C-terminal end of the loop between β2 and β3. The structure shows the metal buried deeply within the protein, an observation that is supported by the inability to remove the metal with excess EDTA at room temperature. Circular dichroism spectroscopy shows that this stability extends to high temperature, with essentially no change being observed in the CD spectrum of Mt-RubB upon heating to 353 K.
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spelling pubmed-31694172011-09-21 Solution-state NMR structure and biophysical characterization of zinc-substituted rubredoxin B (Rv3250c) from Mycobacterium tuberculosis Buchko, Garry W. Hewitt, Stephen N. Napuli, Alberto J. Van Voorhis, Wesley C. Myler, Peter J. Acta Crystallogr Sect F Struct Biol Cryst Commun Structural Communications Owing to the evolution of multi-drug-resistant and extremely drug-resistant Mycobacterium tuberculosis strains, there is an urgent need to develop new antituberculosis strategies to prevent TB epidemics in the industrial world. Among the potential new drug targets are two small nonheme iron-binding proteins, rubredoxin A (Rv3251c) and rubredoxin B (Rv3250c), which are believed to play a role in electron-transfer processes. Here, the solution structure and biophysical properties of one of these two proteins, rubredoxin B (Mt-RubB), determined in the zinc-substituted form are reported. The zinc-substituted protein was prepared by expressing Mt-RubB in minimal medium containing excess zinc acetate. Size-exclusion chromatography and NMR spectroscopy indicated that Mt-RubB was a monomer in solution. The structure (PDB entry 2kn9) was generally similar to those of other rubredoxins, containing a three-stranded anti­parallel β-sheet (β2–β1–β3) and a metal tetrahedrally coordinated to the S atoms of four cysteine residues (Cys9, Cys12, Cys42 and Cys45). The first pair of cysteine residues is at the C-terminal end of the first β-­strand and the second pair of cysteine residues is towards the C-terminal end of the loop between β2 and β3. The structure shows the metal buried deeply within the protein, an observation that is supported by the inability to remove the metal with excess EDTA at room temperature. Circular dichroism spectroscopy shows that this stability extends to high temperature, with essentially no change being observed in the CD spectrum of Mt-RubB upon heating to 353 K. International Union of Crystallography 2011-08-16 /pmc/articles/PMC3169417/ /pubmed/21904065 http://dx.doi.org/10.1107/S1744309111008189 Text en © Buchko et al. 2011 http://creativecommons.org/licenses/by/2.0/uk/ This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.
spellingShingle Structural Communications
Buchko, Garry W.
Hewitt, Stephen N.
Napuli, Alberto J.
Van Voorhis, Wesley C.
Myler, Peter J.
Solution-state NMR structure and biophysical characterization of zinc-substituted rubredoxin B (Rv3250c) from Mycobacterium tuberculosis
title Solution-state NMR structure and biophysical characterization of zinc-substituted rubredoxin B (Rv3250c) from Mycobacterium tuberculosis
title_full Solution-state NMR structure and biophysical characterization of zinc-substituted rubredoxin B (Rv3250c) from Mycobacterium tuberculosis
title_fullStr Solution-state NMR structure and biophysical characterization of zinc-substituted rubredoxin B (Rv3250c) from Mycobacterium tuberculosis
title_full_unstemmed Solution-state NMR structure and biophysical characterization of zinc-substituted rubredoxin B (Rv3250c) from Mycobacterium tuberculosis
title_short Solution-state NMR structure and biophysical characterization of zinc-substituted rubredoxin B (Rv3250c) from Mycobacterium tuberculosis
title_sort solution-state nmr structure and biophysical characterization of zinc-substituted rubredoxin b (rv3250c) from mycobacterium tuberculosis
topic Structural Communications
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3169417/
https://www.ncbi.nlm.nih.gov/pubmed/21904065
http://dx.doi.org/10.1107/S1744309111008189
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