Label-Free Optical Detection of Lysine Acetylation in a Protein using UV-Vis Charge Transfer Signatures

Protein acetylation, an important post-translational modification (PTM) that modulates diverse cellular processes, is presently detected by antibody-based cost-intensive and/or destructive techniques such as mass spectrometry. Here, we show that the novel UV-Vis Protein Charge transfer spectra (ProCharTS) discovered recently by us offers a label-free, easily accessible, and cost-effective option to detect protein acetylation. Specifically, we demonstrate the monotonic decrease of the ProCharTS extinction coefficient between 370-800 nm to increasing degrees of chemical acetylation in two charge-rich proteins α3C and α3W. While our analysis effectively rules out any acetylation induced changes in protein secondary structure, tryptophan fluorescence in α3W and molecular dynamics (MD) simulations hint at the loss of protein compactness with some local unfolding, post-acetylation. We develop a general computational framework based on MD simulations and time-dependent density functional theory (TDDFT) spectra calculations to predict the ProCharTS of whole proteins from their known 3D structure. Computed ProCharTS profiles of acetylated/un-acetylated α3C and α3W enable us to assign the observed PTM induced decrease in intensity to changes in the size, composition and spatial distribution of charged amino acid clusters. Through this experimental and computational approach, we were able to detect 5 or more acetylation events per protein with significant scope for further improvements in sensitivity. More broadly, this study expounds a new optical mode (ProCharTSPTM) exploiting charge transfer transitions to probe/track charged residue modifications in protein solutions.