Structural Studies on the M. tuberculosis Nucleoid-associated-Protein, NapA, Indicates DNA Bridging Mechanism.

Area of Science:

• Bacterial molecular biology
• Structural biology
• Genomics

Background:

• Nucleoid-associated proteins (NAPs) are crucial for bacterial DNA organization.
• Mycobacterium tuberculosis (Mtb) uniquely lacks many common NAPs.
• The essential Mtb NAP, NapA, has unknown DNA-binding functions due to lack of sequence homology.

Purpose of the Study:

• To elucidate the structure and DNA-binding mechanism of the essential Mtb NAP, NapA.
• To understand how NapA organizes the bacterial chromosome.

Main Methods:

• X-ray crystallography to determine apo and DNA-bound NapA structures.
• Mass photometry to analyze NapA oligomerization.
• Biophysical analyses of DNA binding and bridging.

Main Results:

• NapA adopts a dimeric fold, forming higher-order dimer-of-dimers at higher concentrations.
• The dimeric structure features an extended α1 helix and a three-helix module.
• NapA utilizes two distinct DNA-interacting elements per dimer to bind and bridge DNA sites.

Conclusions:

• NapA's unique dimeric structure facilitates DNA binding and bridging, essential for Mtb chromosome organization.
• NapA represents a novel class of DNA-binding proteins conserved in Actinobacteria.
• Structural insights provide a mechanistic understanding of this essential bacterial protein.