Atomic force microscopy studies of APOBEC3G oligomerization and dynamics

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• APOBEC3G (A3G) is a crucial innate immune protein that deaminates DNA and restricts viral replication.
• Its biophysical properties, particularly oligomerization and DNA interaction mechanisms, remain incompletely understood.
• Key questions concern A3G stoichiometry, factors influencing its oligomeric state, and dynamics upon DNA binding.

Purpose of the Study:

• To investigate the oligomerization properties of APOBEC3G (A3G) in free and DNA-bound states.
• To determine the stoichiometry, influencing factors, and dynamics of A3G oligomers.
• To elucidate the relationship between A3G oligomer size and single-stranded DNA (ssDNA) substrate length.

Main Methods:

• Utilized atomic force microscopy (AFM) for direct imaging of A3G protein.
• Employed time-lapse AFM imaging to characterize the dynamics of A3G oligomers.
• Analyzed A3G in both free states and in complexes with single-stranded DNA (ssDNA).

Main Results:

• A3G inherently forms oligomers, with formation yield dependent on protein concentration.
• Oligomer size increases with higher protein concentrations, both in free states and when bound to ssDNA.
• A3G oligomers spontaneously dissociate into monomers, primarily via monomer dissociation.

Conclusions:

• Oligomerization is an intrinsic property of A3G, modulated by protein concentration.
• A3G binding to ssDNA shows size-dependent substrate length requirements, with larger oligomers needing longer DNA.
• The dissociation of A3G oligomers occurs spontaneously, mainly through monomer release.