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Published on: February 25, 2010
Exopolyphosphatases PPX1 and PPX2 from Corynebacterium glutamicum
Steffen N Lindner1, Sandra Knebel, Hendrik Wesseling
1Institut für Biotechnologie 1, Forschungszentrum Jülich, D-52425 Jülich, Germany.
Applied and Environmental Microbiology
|March 24, 2009
Summary
Corynebacterium glutamicum utilizes two exopolyphosphatases (PPX1 and PPX2) to regulate inorganic polyphosphate (PolyP) levels. PPX2 is identified as the primary enzyme, crucial for growth under phosphate limitation.
Area of Science:
- Microbiology
- Biochemistry
- Molecular Biology
Background:
- Corynebacterium glutamicum accumulates significant intracellular inorganic polyphosphate (PolyP).
- PolyP serves as a phosphate reservoir and influences cellular processes.
- The enzymes responsible for PolyP metabolism in C. glutamicum were not fully characterized.
Purpose of the Study:
- To identify and characterize the exopolyphosphatases (PPX) in Corynebacterium glutamicum.
- To investigate the roles of PPX1 and PPX2 in PolyP metabolism and cellular growth.
- To determine the enzymatic properties of the major exopolyphosphatase.
Main Methods:
- Gene expression analysis (overexpression and deletion mutants) of cg0488 (ppx1) and cg1115 (ppx2).
- Measurement of exopolyphosphatase activity in crude extracts and purified enzyme.
- Quantification of intracellular PolyP concentrations under various conditions.
- Growth experiments comparing wild-type, mutant, and overexpression strains.
- Enzyme purification and characterization of PPX2(His).
Main Results:
- Both PPX1 and PPX2 exhibit exopolyphosphatase activity, with PPX2 being the major contributor.
- Deletion mutants showed increased intracellular PolyP, while overexpression strains had decreased PolyP.
- Growth advantage was observed for deletion mutants under phosphate-limiting conditions, and disadvantage for overexpression strains.
- Purified PPX2 functions as a monomer, requires Mg2+ or Mn2+, and is inhibited by millimolar divalent cations and nucleoside triphosphates.
- PPX2 efficiently hydrolyzes short-chain polyphosphates and pyrophosphate.
Conclusions:
- PPX2 is the principal exopolyphosphatase in C. glutamicum, playing a key role in inorganic polyphosphate homeostasis.
- Differential regulation of PPX1 and PPX2 impacts intracellular PolyP levels and cellular adaptation to phosphate availability.
- Understanding PPX enzymes provides insights into microbial phosphate metabolism and stress response.
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