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Structure-Function Analysis of the Transmembrane Protein AmpG from Pseudomonas aeruginosa

AmpG is a transmembrane protein with permease activity that transports meuropeptide from the periplasm to the cytoplasm, which is essential for the induction of the ampC encoding β-lactamase. To obtain new insights into the relationship between AmpG structure and function, comparative genomics analy...

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Autores principales: Li, Peizhen, Ying, Jun, Yang, Guangjian, Li, Aifang, Wang, Jian, Lu, Junwan, Wang, Junrong, Xu, Teng, Yi, Huiguang, Li, Kewei, Jin, Shouguang, Bao, Qiyu, Zhang, Kaibo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5154545/
https://www.ncbi.nlm.nih.gov/pubmed/27959942
http://dx.doi.org/10.1371/journal.pone.0168060
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author Li, Peizhen
Ying, Jun
Yang, Guangjian
Li, Aifang
Wang, Jian
Lu, Junwan
Wang, Junrong
Xu, Teng
Yi, Huiguang
Li, Kewei
Jin, Shouguang
Bao, Qiyu
Zhang, Kaibo
author_facet Li, Peizhen
Ying, Jun
Yang, Guangjian
Li, Aifang
Wang, Jian
Lu, Junwan
Wang, Junrong
Xu, Teng
Yi, Huiguang
Li, Kewei
Jin, Shouguang
Bao, Qiyu
Zhang, Kaibo
author_sort Li, Peizhen
collection PubMed
description AmpG is a transmembrane protein with permease activity that transports meuropeptide from the periplasm to the cytoplasm, which is essential for the induction of the ampC encoding β-lactamase. To obtain new insights into the relationship between AmpG structure and function, comparative genomics analysis, secondary and tertiary structure modeling, site-directed mutational analyses and genetic complementation experiments were performed in this study. AmpGs from different genera of bacteria (Escherichia coli, Vibrio cholerae and Acinetobacter baumannii) could complement AmpG function in Pseudomonas aeruginosa. The minimal inhibitory concentration (MIC) to ampicillin is 512 μg/ml for wild type strain PAO1, while it is 32 μg/ml for an ampG deletion mutant strain (PAO1ΔampG) with a corresponding decrease in the activity of the ampC-encoded β-lactamase. Site-directed mutagenesis of conserved AmpG residues (G29, A129, Q131 and A197) resulted in a loss of function, resulting in a loss of resistance to ampicillin in PAO1ΔampG. The G29A, G29V, A129T, A129V, A129D, A197S and A197D mutants had lower resistance to ampicillin and significantly decreased activity of the AmpC β-lactamase. The G29A, G29V, A129V, A197S and A197D mutants had decreased ampG mRNA transcript levels. The A129T and A129D mutants had normal ampG mRNA transcript levels, but the function of the protein was drastically reduced. Our experimental results demonstrate that the conserved amino acids played essential roles in maintaining the function of AmpG. Combined with the AmpG structural information, these critical amino acids can be targeted for the development of new anti-bacterial agents.
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spelling pubmed-51545452016-12-28 Structure-Function Analysis of the Transmembrane Protein AmpG from Pseudomonas aeruginosa Li, Peizhen Ying, Jun Yang, Guangjian Li, Aifang Wang, Jian Lu, Junwan Wang, Junrong Xu, Teng Yi, Huiguang Li, Kewei Jin, Shouguang Bao, Qiyu Zhang, Kaibo PLoS One Research Article AmpG is a transmembrane protein with permease activity that transports meuropeptide from the periplasm to the cytoplasm, which is essential for the induction of the ampC encoding β-lactamase. To obtain new insights into the relationship between AmpG structure and function, comparative genomics analysis, secondary and tertiary structure modeling, site-directed mutational analyses and genetic complementation experiments were performed in this study. AmpGs from different genera of bacteria (Escherichia coli, Vibrio cholerae and Acinetobacter baumannii) could complement AmpG function in Pseudomonas aeruginosa. The minimal inhibitory concentration (MIC) to ampicillin is 512 μg/ml for wild type strain PAO1, while it is 32 μg/ml for an ampG deletion mutant strain (PAO1ΔampG) with a corresponding decrease in the activity of the ampC-encoded β-lactamase. Site-directed mutagenesis of conserved AmpG residues (G29, A129, Q131 and A197) resulted in a loss of function, resulting in a loss of resistance to ampicillin in PAO1ΔampG. The G29A, G29V, A129T, A129V, A129D, A197S and A197D mutants had lower resistance to ampicillin and significantly decreased activity of the AmpC β-lactamase. The G29A, G29V, A129V, A197S and A197D mutants had decreased ampG mRNA transcript levels. The A129T and A129D mutants had normal ampG mRNA transcript levels, but the function of the protein was drastically reduced. Our experimental results demonstrate that the conserved amino acids played essential roles in maintaining the function of AmpG. Combined with the AmpG structural information, these critical amino acids can be targeted for the development of new anti-bacterial agents. Public Library of Science 2016-12-13 /pmc/articles/PMC5154545/ /pubmed/27959942 http://dx.doi.org/10.1371/journal.pone.0168060 Text en © 2016 Li et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Li, Peizhen
Ying, Jun
Yang, Guangjian
Li, Aifang
Wang, Jian
Lu, Junwan
Wang, Junrong
Xu, Teng
Yi, Huiguang
Li, Kewei
Jin, Shouguang
Bao, Qiyu
Zhang, Kaibo
Structure-Function Analysis of the Transmembrane Protein AmpG from Pseudomonas aeruginosa
title Structure-Function Analysis of the Transmembrane Protein AmpG from Pseudomonas aeruginosa
title_full Structure-Function Analysis of the Transmembrane Protein AmpG from Pseudomonas aeruginosa
title_fullStr Structure-Function Analysis of the Transmembrane Protein AmpG from Pseudomonas aeruginosa
title_full_unstemmed Structure-Function Analysis of the Transmembrane Protein AmpG from Pseudomonas aeruginosa
title_short Structure-Function Analysis of the Transmembrane Protein AmpG from Pseudomonas aeruginosa
title_sort structure-function analysis of the transmembrane protein ampg from pseudomonas aeruginosa
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5154545/
https://www.ncbi.nlm.nih.gov/pubmed/27959942
http://dx.doi.org/10.1371/journal.pone.0168060
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