Crystallization And Structure Determination Of The Phycobilisome Complex
Written by Liron David
Liron David and Noam Adir
Schulich Faculty of Chemistry, Technion, Israel Institute of Technology, Haifa, Israel
The photosynthetic process is initiated by the absorption of light energy by protein complexes called light harvesting antennas. In cyanobacteria and red algae the major antenna is called the Phycobilisome (PBS).
The PBS is an extremely large complex, with a molecular weight of 3-7MDa which is made up of pigmented proteins known as phycobiliproteins (PBPs) and unpigmented proteins known as linker proteins.Our goal is to obtain an atomic resolution structure of the entire PBS complex from the cyanobacterium Thermosynechococcus vulcanus using x-ray crystallography. Intact PBS was isolated in high phosphate buffer by sucrose gradient ultracentrifugation. Small blue crystals shaped like half moons were obtained in stabilization buffer in two to four weeks. Material obtained from the dissolving of extensively washed crystals was analyzed by fluorescence, SDS-PAGE and mass spectrometry (MS). The results of these experiments indicate that the crystals contain intact, functional PBS complex. Dynamic light scattering indicates a molecular weight of at least 2.8MDa. Preliminary diffraction experiments have indicated that the present crystals diffract to 3.5Å.A structure of phycocyanin (one of the PBP components) was determined as a half hexamer in the asymmetric unit. Within the phycocyanin disks, unstructured electron density could be identified in the position thought to be occupied by the linker. MS results for those crystals suggest that this is a phycocyanin rod structure contain the phycocyanin and three different linker proteins.