Non-invasive, ratiometric determination of intracellular pH in Pseudomonas species using a novel genetically encoded indicator

Alejandro Arce-Rodríguez, Daniel Christoph Volke, Sarina Bense, Susanne Häussler, Pablo Ivan Nikel

Research output: Contribution to journalJournal articleResearchpeer-review

69 Downloads (Pure)

Abstract

The ability of Pseudomonas species to thrive in all major natural environments (i.e. terrestrial, freshwater and marine) is based on its exceptional capability to adapt to physicochemical changes. Thus, environmental bacteria have to tightly control the maintenance of numerous physiological traits across different conditions. The intracellular pH (pHi) homoeostasis is a particularly important feature, since the pHi influences a large portion of the biochemical processes in the cell. Despite its importance, relatively few reliable, easy-to-implement tools have been designed for quantifying in vivo pHi changes in Gram-negative bacteria with minimal manipulations. Here we describe a convenient, non-invasive protocol for the quantification of the pHi in bacteria, which is based on the ratiometric fluorescent indicator protein PHP (pH indicator for Pseudomonas). The DNA sequence encoding PHP was thoroughly adapted to guarantee optimal transcription and translation of the indicator in Pseudomonas species. Our PHP-based quantification method demonstrated that pHi is tightly regulated over a narrow range of pH values not only in Pseudomonas, but also in other Gram-negative bacterial species such as Escherichia coli. The maintenance of the cytoplasmic pH homoeostasis in vivo could also be observed upon internal (e.g. redirection of glucose consumption pathways in P. putida) and external (e.g. antibiotic exposure in P. aeruginosa) perturbations, and the PHP indicator was also used to follow dynamic changes in the pHi upon external pH shifts. In summary, our work describes a reliable method for measuring pHi in Pseudomonas, allowing for the detailed investigation of bacterial pHi homoeostasis and its regulation.
Original languageEnglish
JournalMicrobial Biotechnology
Volume12
Issue number4
Pages (from-to)799-813
DOIs
Publication statusPublished - 2019

Keywords

  • Biotechnology
  • Bioengineering
  • Biochemistry
  • Applied Microbiology and Biotechnology

Cite this

@article{6f8555d4c7094db1bf8cad4eeabe234c,
title = "Non-invasive, ratiometric determination of intracellular pH in Pseudomonas species using a novel genetically encoded indicator",
abstract = "The ability of Pseudomonas species to thrive in all major natural environments (i.e. terrestrial, freshwater and marine) is based on its exceptional capability to adapt to physicochemical changes. Thus, environmental bacteria have to tightly control the maintenance of numerous physiological traits across different conditions. The intracellular pH (pHi) homoeostasis is a particularly important feature, since the pHi influences a large portion of the biochemical processes in the cell. Despite its importance, relatively few reliable, easy-to-implement tools have been designed for quantifying in vivo pHi changes in Gram-negative bacteria with minimal manipulations. Here we describe a convenient, non-invasive protocol for the quantification of the pHi in bacteria, which is based on the ratiometric fluorescent indicator protein PHP (pH indicator for Pseudomonas). The DNA sequence encoding PHP was thoroughly adapted to guarantee optimal transcription and translation of the indicator in Pseudomonas species. Our PHP-based quantification method demonstrated that pHi is tightly regulated over a narrow range of pH values not only in Pseudomonas, but also in other Gram-negative bacterial species such as Escherichia coli. The maintenance of the cytoplasmic pH homoeostasis in vivo could also be observed upon internal (e.g. redirection of glucose consumption pathways in P. putida) and external (e.g. antibiotic exposure in P. aeruginosa) perturbations, and the PHP indicator was also used to follow dynamic changes in the pHi upon external pH shifts. In summary, our work describes a reliable method for measuring pHi in Pseudomonas, allowing for the detailed investigation of bacterial pHi homoeostasis and its regulation.",
keywords = "Biotechnology, Bioengineering, Biochemistry, Applied Microbiology and Biotechnology",
author = "Alejandro Arce-Rodr{\'i}guez and Volke, {Daniel Christoph} and Sarina Bense and Susanne H{\"a}ussler and Nikel, {Pablo Ivan}",
year = "2019",
doi = "10.1111/1751-7915.13439",
language = "English",
volume = "12",
pages = "799--813",
journal = "Microbial Biotechnology",
issn = "1751-7915",
publisher = "Wiley-Blackwell",
number = "4",

}

Non-invasive, ratiometric determination of intracellular pH in Pseudomonas species using a novel genetically encoded indicator. / Arce-Rodríguez, Alejandro; Volke, Daniel Christoph; Bense, Sarina; Häussler, Susanne; Nikel, Pablo Ivan.

In: Microbial Biotechnology, Vol. 12, No. 4, 2019, p. 799-813.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Non-invasive, ratiometric determination of intracellular pH in Pseudomonas species using a novel genetically encoded indicator

AU - Arce-Rodríguez, Alejandro

AU - Volke, Daniel Christoph

AU - Bense, Sarina

AU - Häussler, Susanne

AU - Nikel, Pablo Ivan

PY - 2019

Y1 - 2019

N2 - The ability of Pseudomonas species to thrive in all major natural environments (i.e. terrestrial, freshwater and marine) is based on its exceptional capability to adapt to physicochemical changes. Thus, environmental bacteria have to tightly control the maintenance of numerous physiological traits across different conditions. The intracellular pH (pHi) homoeostasis is a particularly important feature, since the pHi influences a large portion of the biochemical processes in the cell. Despite its importance, relatively few reliable, easy-to-implement tools have been designed for quantifying in vivo pHi changes in Gram-negative bacteria with minimal manipulations. Here we describe a convenient, non-invasive protocol for the quantification of the pHi in bacteria, which is based on the ratiometric fluorescent indicator protein PHP (pH indicator for Pseudomonas). The DNA sequence encoding PHP was thoroughly adapted to guarantee optimal transcription and translation of the indicator in Pseudomonas species. Our PHP-based quantification method demonstrated that pHi is tightly regulated over a narrow range of pH values not only in Pseudomonas, but also in other Gram-negative bacterial species such as Escherichia coli. The maintenance of the cytoplasmic pH homoeostasis in vivo could also be observed upon internal (e.g. redirection of glucose consumption pathways in P. putida) and external (e.g. antibiotic exposure in P. aeruginosa) perturbations, and the PHP indicator was also used to follow dynamic changes in the pHi upon external pH shifts. In summary, our work describes a reliable method for measuring pHi in Pseudomonas, allowing for the detailed investigation of bacterial pHi homoeostasis and its regulation.

AB - The ability of Pseudomonas species to thrive in all major natural environments (i.e. terrestrial, freshwater and marine) is based on its exceptional capability to adapt to physicochemical changes. Thus, environmental bacteria have to tightly control the maintenance of numerous physiological traits across different conditions. The intracellular pH (pHi) homoeostasis is a particularly important feature, since the pHi influences a large portion of the biochemical processes in the cell. Despite its importance, relatively few reliable, easy-to-implement tools have been designed for quantifying in vivo pHi changes in Gram-negative bacteria with minimal manipulations. Here we describe a convenient, non-invasive protocol for the quantification of the pHi in bacteria, which is based on the ratiometric fluorescent indicator protein PHP (pH indicator for Pseudomonas). The DNA sequence encoding PHP was thoroughly adapted to guarantee optimal transcription and translation of the indicator in Pseudomonas species. Our PHP-based quantification method demonstrated that pHi is tightly regulated over a narrow range of pH values not only in Pseudomonas, but also in other Gram-negative bacterial species such as Escherichia coli. The maintenance of the cytoplasmic pH homoeostasis in vivo could also be observed upon internal (e.g. redirection of glucose consumption pathways in P. putida) and external (e.g. antibiotic exposure in P. aeruginosa) perturbations, and the PHP indicator was also used to follow dynamic changes in the pHi upon external pH shifts. In summary, our work describes a reliable method for measuring pHi in Pseudomonas, allowing for the detailed investigation of bacterial pHi homoeostasis and its regulation.

KW - Biotechnology

KW - Bioengineering

KW - Biochemistry

KW - Applied Microbiology and Biotechnology

U2 - 10.1111/1751-7915.13439

DO - 10.1111/1751-7915.13439

M3 - Journal article

VL - 12

SP - 799

EP - 813

JO - Microbial Biotechnology

JF - Microbial Biotechnology

SN - 1751-7915

IS - 4

ER -