Protein

Protein

Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...

 footprinting involves the study of the surface of proteins to investigate protein structure

Protein structure

Proteins are an important class of biological macromolecules present in all organisms. Proteins are polymers of amino acids. Classified by their physical size, proteins are nanoparticles . Each protein polymer – also known as a polypeptide – consists of a sequence formed from 20 possible L-α-amino...

 or how they assemble and interact within a larger macromolecular assembly. This has been traditionally achieved through the treatment of proteins with hydroxyl radicals.

Time-resolved hydroxyl radical protein footprinting employing mass spectrometry analysis was developed over a decade ago in synchrotron

Synchrotron

A synchrotron is a particular type of cyclic particle accelerator in which the magnetic field and the electric field are carefully synchronised with the travelling particle beam. The proton synchrotron was originally conceived by Sir Marcus Oliphant...

 radiolysis studies conduced by Maleknia et al.. The same year, these authors reported on the use of an electrical discharge source to effect the oxidation of proteins on millisecond timescales as proteins pass from the electrosprayed solution into the mass spectrometer These approaches have since been successfully applied in the analysis of protein structures, protein folding, protein dynamics, and protein–protein interactions.

Method

The exposure of proteins to a ‘white’ X-ray beam of synchrotron light or an electrical discharge for tens of milliseconds provides sufficient oxidative modification to the surface amino acid side chains without damage to the protein structure. These products can be easily detected and quantified by mass spectrometry (Figure 1). By adjusting the time for radiolysis or which protein ions spend in the discharge source, a time-resolved approach is possible which is valuable for the study of protein dynamics.