Laser‐Initiated Radical Trifluoromethylation of Peptides and Proteins: Application to Mass‐Spectrometry‐Based Protein Footprinting

Abstract

We describe a novel, laser-initiated radical trifluoromethylation for protein footprinting and establish its broad residue coverage. •CF3 reacts with 18 of 20 common amino acids including Gly, Ala, Ser, Thr, Asp, Glu that are relatively “silent” with •OH. This new approach to footprinting is a bridge between trifluoromethylation in materials and medicinal chemistry and structural biology and biotechnology. Its application to a membrane protein and to myoglobin show that the approach is sensitive to protein conformational change and solvent accessibility.

The Application of Fluorine‐Containing Reagents in Structural Proteomics

“…Recently, Gross and co‐workers capitalized on the trifluoromethyl radical ( . CF 3 ) as a footprinting reagent generated by pulsed laser photolysis on the FPOP platform (Figure ).…”

“…Even with quenchers such as DMSO or detergents, which are often in protein solutions, . CF 3 maintains its reactivity towards proteins and small organic molecules The Langlois reagent, however, cannot be directly photolyzed to give the radical.…”

Fluorierte Reagenzien in der Strukturproteomik

“…Vor kurzem haben Gross und Mitarbeiter das Trifluormethylradikal ( . CF 3 ) als Footprinting‐Reagenz eingeführt, das durch pulsierende Laser‐Photolyse auf der FPOP‐Plattform erzeugt wird (Abbildung ) . Das hochreaktive .…”

“…Sogar in Gegenwart von Quenchern wie DMSO oder Detergenzien, die oft in Proteinlösungen gefunden werden, behält . CF 3 seine Reaktivität mit Proteinen und niedermolekularen organischen Verbindungen Die .…”

In Vivo Hydroxyl Radical Protein Footprinting for the Study of Protein Interactions in <em>Caenorhabditis elegans</em>

“…These techniques can bias towards one protein conformer if the epitope/binding site of the protein has been oxidatively modified by IV-FPOP. New developments in footprinting radical reagents, such as sulfate radical anion 10 or trifluoromethylation 19 could increase the versatility of IV-FPOP. Although the only labeling reagent tested in vivo thus far is hydrogen peroxide, other laser-activated radicals could be optimized.…”