Aggregation-related quenching of LHCII fluorescence in liposomes revealed by single-molecule spectroscopy

Incorporation of membrane proteins into reconstituted lipid membranes is a common approach for studying their structure and function relationship in a native-like environment. In this work, we investigated fluorescence properties of liposome-reconstituted major light-harvesting complexes of plants (LHCII). By utilizing liposome labelling with the fluorescent dye molecules and single-molecule microscopy techniques, we were able to study truly liposome-reconstituted LHCII and compare them with bulk measurements and liposome-free LHCII aggregates bound to the surface. Our results showed that fluorescence lifetime obtained in bulk and in single liposome measurements were correlated. The fluorescence lifetimes of LHCII were shorter for liposome-free LHCII than for reconstituted LHCII. In the case of liposome-reconstituted LHCII, fluorescence lifetime showed dependence on the protein density reminiscent to concentration quenching. The dependence of fluorescence lifetime of LHCII on the liposome size was not significant. Our results demonstrated that fluorescence quenching can be induced by LHCII - LHCII interactions in reconstituted membranes, most likely occurring via the same mechanism as photoprotective non-photochemical quenching in vivo.

Automated summary

This study investigated the fluorescence properties of liposome-reconstituted major light-harvesting complexes of plants (LHCII) using fluorescent dye labelling and single-molecule microscopy techniques. Results showed that the fluorescence lifetimes in bulk and single liposome measurements were correlated, with shorter lifetimes observed in liposome-free LHCII compared to reconstituted LHCII. The dependence of fluorescence lifetime on protein density in liposome-reconstituted LHCII resembled concentration quenching, while the impact of liposome size on fluorescence lifetime was not significant. It was demonstrated that fluorescence quenching in reconstituted membranes could be induced by LHCII-LHCII interactions, similar to photoprotective non-photochemical quenching in vivo.