Photoinduced decarboxylation in fluorescent proteins: charge-transfer states and structure-function relationship

Photoinduced decarboxylation of acidic amino acids near the chromophore is observed in various fluorescent protein variants, including avGFP, PA-GFP, DsRed, LSSmOrange, and PSLSSmKate. While a mechanism has been proposed and theoretically supported for GFPs, its generalization to other variants remains unclear. This study investigates the decarboxylation mechanism across different fluorescent proteins, with a particular focus on the red fluorescent proteins DsRed and PSLSSmKate, to establish a structure-function relationship. A benchmark study was conducted to assess various excited-state methods, basis sets, and embedding approaches. Excited-state QM/MM calculations using TD-DFT, ADC(2), and CC2 methods were employed to explore the excited states of multiple fluorescent protein variants. Our findings provide new insights into the decarboxylation mechanism in fluorescent proteins, contributing to the broader field of protein photophysics and potentially informing the design of novel fluorescent probes.