Channelrhodopsins (ChRs) are light-gated ion channels that control photomovement of microalgae. In optogenetics, ChRs are widely applied for light-triggering action potentials in cells, tissues, and living animals, yet the spectral properties and photocycle of ChR remain obscure. In this study, we cloned a ChR from the colonial alga Volvox carteri, VChR. After electrophysiological characterization in Xenopus oocytes, VChR was expressed in COS-1 cells and purified. Time-resolved UV-visible spectroscopy revealed a pH-dependent equilibrium of two dark species, D(470)/D(480). Laser flashes converted both with tau approximately 200 mus into major photointermediates P(510)/P(530), which reverted back to the dark states with tau approximately 15-100 ms. Both intermediates were assigned to conducting states. Three early intermediates P(500)/P(515) and P(390) were detected on a ns to mus time scale. The spectroscopic and electrical data were unified in a photocycle model. The functional expression of VChR we report here paves the way toward a broader structure/function analysis of the recently identified class of light-gated ion channels.

EEF1A2 is one of the isoforms of the alpha subunit of the eukaryotic Elongation Factor 1. It is overexpressed in human tumors and is endowed with oncogenic properties, favoring tumor cell proliferation while inhibiting apoptosis. We demonstrate that plitidepsin, an antitumor agent of marine origin that has successfully completed a phase-III clinical trial for multiple myeloma, exerts its antitumor activity by targeting eEF1A2. The drug interacts with eEF1A2 with aK_Dof 80 nM and a target residence time of circa 9 min. This protein was also identified as capable of binding [¹⁴C]-plitidepsin in a cell lysate from K-562 tumor cells. A molecular modelling approach was used to identify a favorable binding site for plitidepsin at the interface between domains 1 and 2 of eEF1A2 in the GTP conformation. Three tumor cell lines selected for at least 100-fold more resistance to plitidepsin than their respective parental cells showed reduced levels of eEF1A2 protein. Ectopic expression of eEF1A2 in resistant cells restored the sensitivity to plitidepsin. FLIM-phasor FRET experiments demonstrated that plitidepsin localizes in tumor cells sufficiently close to eEF1A2 as to suggest the formation of drug-protein complexes in living cells. Altogether, our results strongly suggest that eEF1A2 is the primary target of plitidepsin.

The ultrafast time evolution of a single-stranded adenine DNA is studied using an hybrid multiscale quantum mechanics/molecular mechanics (QM/MM) scheme coupled to nonadiabatic surface hopping dynamics. As a model we use (dA)$_{20}$ where a stacked adenine tetramer is treated quantum chemically. The dynamical simulations combined with on-the-fly quantitative wavefunction analysis evidence the nature of the long-lived electronically excited states formed upon absorption of UV light. After a rapid decrease of the initially excited excitons, relaxation to monomer-like states and excimers occurs within 100 fs The former monomeric states then relax into additional excimer states en route to form stabilised charge transfer states on a longer time-scale of hundreds of fs. The different electronic state characters manifest on the spatial separation between the adenines: excimers and charge transfer states show a much smaller spatial separation than the monomer-like states and the initially formed excitons.