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Crystallization of a stringent response factor from Aquifex aeolicus
Ole Kristensen1, Martin Laurberg, Michael Gajhede
1Protein Structure Group, Department of Chemistry, University of Copenhagen, Universitetsparken 5, Denmark. olek@psg.ki.ku.dk
Summary
The crystallization of a bifunctional enzyme from Aquifex aeolicus, acting as an exopolyphosphatase and guanosine pentaphosphate phosphohydrolase, was achieved. This study reports the first crystallization of a Ppx/GppA family phosphatase member.
Area of Science:
- Biochemistry
- Structural Biology
- Enzymology
Background:
- Aquifex aeolicus harbors enzymes with potential bifunctional catalytic activity.
- The Ppx/GppA phosphatase family plays crucial roles in cellular metabolism.
- Understanding the structure of these enzymes is key to elucidating their function.
Purpose of the Study:
- To report the successful crystallization of a key enzyme from Aquifex aeolicus.
- To characterize the enzyme's bifunctional activity as an exopolyphosphatase and guanosine pentaphosphate phosphohydrolase.
- To obtain high-resolution structural data for future mechanistic studies.
Main Methods:
- Crystallization of the native enzyme from Aquifex aeolicus.
- X-ray diffraction data collection to below 2 Å resolution.
- Mutagenesis to introduce methionine residues and protein oxidation to generate new crystal forms.
Main Results:
- An orthorhombic crystal form with specific unit-cell parameters was obtained.
- Native data resolution was achieved below 2 Å.
- Multiple crystal forms with improved diffraction and phasing potential were generated through protein modification.
Conclusions:
- This work represents the first reported crystallization of a Ppx/GppA family phosphatase.
- The obtained crystal forms provide a foundation for high-resolution structural determination.
- Further structural studies will aid in understanding the enzyme's bifunctional catalytic mechanism.