Dissecting pigment architecture of individual photosynthetic antenna complexes in solution.

Oligomerization plays a critical role in shaping the light-harvesting properties of many photosynthetic pigment-protein complexes, but a detailed understanding of this process at the level of individual pigments is still lacking. To study the effects of oligomerization, we designed a single-molecule approach to probe the photophysical properties of individual pigment sites as a function of protein assembly state. Our method, based on the principles of anti-Brownian electrokinetic trapping of single fluorescent proteins, step-wise photobleaching, and multiparameter spectroscopy, allows pigment-specific spectroscopic information on single multipigment antennae to be recorded in a nonperturbative aqueous environment with unprecedented detail. We focus on the monomer-to-trimer transformation of allophycocyanin (APC), an important antenna protein in cyanobacteria. Our data reveal that the two chemically identical pigments in APC have different roles. One (α) is the functional pigment that red-shifts its spectral properties upon trimer formation, whereas the other (β) is a "protective" pigment that persistently quenches the excited state of α in the prefunctional, monomer state of the protein. These results show how subtleties in pigment organization give rise to functionally important aspects of energy transfer and photoprotection in antenna complexes. The method developed here should find immediate application in understanding the emergent properties of other natural and artificial light-harvesting systems.