Functional phycobilisome core structures in a phycocyanin-less mutant of cyanobacterium Synechococcus sp. PCC 7942

Abstract

We have constructed a mutant Synechococcus sp. PCC 7942, termed R2HECAT, in which the entire phycobilisome rod operon has been deleted. In the whole cell absorption spectra of R2HECAT, the peak corresponding to phycocyanin (PC), λmax≈620 nm, could not be detected. However, a single pigment-protein fraction with λmax=654 nm could be isolated on sucrose gradients from R2HECAT. Analysis of this pigment-protein fraction by non-denaturing PAGE indicates an apparent molecular mass of about 1200-1300 kDa. On exposure to low temperature, the isolated pigment-protein complex dissociated to a protein complex with a molecular mass of about 560 kDa. When analysed by SDS-PAGE, the pigment-protein fraction was found to consist of the core polypeptides but lacked PC, 27, 33, 30, and the 9 kDa polypeptides which are a part of the rods. All the chromophore bearing polypeptides of the core were found to be chromophorylated. CD as well as absorption spectra showed the expected maxima around 652 and 675 nm from allophycocyanin (APC) and allophycocyanin B (APC-B) chromophores. Low temperature fluorescence and excitation spectra also showed that the core particles were fully functional with respect to the energy transfer between the APC chromophores. We conclude that PC and therefore the rods are dispensable for the survival of Synechococcus sp. PCC 7942. The results indicate that stable and functional core can assemble in absence of the rods. These rod-less phycobilisome core is able to transfer energy to Photosystem II.

Diffusion of Phycobilisomes on the Thylakoid Membranes of the Cyanobacterium Synechococcus 7942

“…The availability of mutants allows us to probe the factors affecting the mobility of the phycobilisomes. We find that phycobilisomes diffuse slightly faster in a mutant in which the phycobilisome rod elements are missing (19), indicating that the size of the phycobilisomes plays a role in determining their rate of diffusion. The temperature dependence of the phycobilisome diffusion coefficient suggests that the phycobilisomes have no integral membrane domain.…”

“…PCC7942 was grown in BG11 medium (21) supplemented with 10 mM NaHCO 3 . For the R2HECAT mutant (19), the growth medium was supplemented with chloramphenicol at 5 g/ml, and for the desA ϩ transformant (20), the medium was supplemented with kanamycin at 50 g/ml. Liquid cultures were grown in an orbital shaking incubator at 30°C with white illumination at approximately 10 microeinsteins m Ϫ2 s Ϫ1 .…”

“…The mutant R2HECAT (19) lacks the cpcA, cpcB, cpcD, cpcE, cpcF, cpcH, and cpcI genes coding for the ␣-and ␤-subunits of phycocyanin and rod-linker polypeptides (19). However, the phycobilisome cores are still assembled and functional (19).…”

“…The mutant R2HECAT (19) lacks the cpcA, cpcB, cpcD, cpcE, cpcF, cpcH, and cpcI genes coding for the ␣-and ␤-subunits of phycocyanin and rod-linker polypeptides (19). However, the phycobilisome cores are still assembled and functional (19). The phycobilisome cores have a molecular mass of 1200 -1300 kDa (19) and dimensions of approximately 22 ϫ 11 ϫ 12 nm (23).…”

“…However, the phycobilisome cores are still assembled and functional (19). The phycobilisome cores have a molecular mass of 1200 -1300 kDa (19) and dimensions of approximately 22 ϫ 11 ϫ 12 nm (23). The intact phycobilisomes of wild-type cells are hemidiscoidal structures with a typical molecular mass of approximately 6000 kDa and typically a longest diameter of approximately 60 nm (23).…”