Highly-Active Recombinant Formate Dehydrogenase from Pathogenic Bacterium Staphylococcus aureus: Preparation and...

Area of Science:

• Biochemistry
• Structural Biology
• Microbiology

Background:

• Staphylococcus aureus formate dehydrogenase (SauFDH) is vital for pathogen survival in biofilms.
• Understanding SauFDH structure can guide the development of specific inhibitors for antibacterial agents.
• Targeting S. aureus biofilms is crucial for combating infections.

Purpose of the Study:

• To produce recombinant SauFDH in high yield.
• To determine the three-dimensional structure of SauFDH.
• To obtain crystals for X-ray analysis.

Main Methods:

• Recombinant expression of SauFDH in E. coli.
• Enzyme purification and activity assays.
• Crystallization and X-ray diffraction analysis using synchrotron sources.

Main Results:

• High yield (1 g/L) and purity (61% yield, 400 mg) of recombinant SauFDH achieved.
• Specific activity of purified SauFDH was 20 U/mg, twice that of previously reported enzymes.
• Successfully obtained crystals and solved the 3D structures of apo- and holo-SauFDH at 2.2 and 2.7 Å resolution, respectively.

Conclusions:

• The study provides the 3D structures of SauFDH apo- and holoenzymes.
• Different crystal space groups for apo- and holo-SauFDH suggest coenzyme binding influences structure.
• The structural data can facilitate the design of novel antibacterial agents targeting S. aureus biofilms.