Candidate Genes for the Phycoerythrocyanin α Subunit Lyase. (1995)

The rod substructures of the Anabaena sp. PCC 7120 phycobilisome contain the light harvesting proteins C-phycocyanin and phycoerythrocyanin (PEC). Even at low light intensities, PEC represents no more than 5% of the phycobilisome protein. The β subunits of both proteins carry thioether-linked phycocyanobilin (PCB) at β-Cys-82 and β-Cys-155; however, C-phycocyanin has PCB at α-Cys-84 whereas PEC α subunit carries phycobiliviolin at this position. The Anabaena sp. PCC 7120 pec operon is made up of five genes. PecB and pecA encode the β and α subunits of PEC, pecC encodes a linker polypeptide associated with PEC in the rod substructure, and pecE and pecF are genes of unknown function that show a high degree of homology to cpcE and cpcF, that encode a C-phycocyanin α subunit PCB lyase (Fairchild, C. D., Zhao, J., Zhou, J., Colson, S. E., Bryant, D. A., and Glazer, A. N.(1992) Proc. Natl. Acad. Sci. U. S. A. 89, 7017-7021). Insertional mutants in pecE and pecF, and an interposon mutant in which a portion of both pecE and pecF was deleted, were constructed. All three types of mutants grew 1.3 times slower than wild-type under limiting light conditions and showed a 20% reduction in the PCB content of whole cells relative to chlorophyll a. Holo-PEC was missing from the phycobilisomes of all three types of mutants and the level of the PEC linker polypeptide was reduced relative to the wild-type. However, ∼30% of the wild-type level of the PEC β subunit was present in all of these phycobilisomes. In contrast, the PEC α subunit was barely detectable in the pecE and pecF mutants, but was present in the pecEF deletion mutant as a PCB-adduct in a 1:1 ratio with the PEC β subunit. The identity of this “unnatural” adduct was confirmed by isolation of the subunit and amino-terminal sequencing. These biochemical results support the inference that pecE and pecF encode a PEC α subunit phycobiliviolin lyase, and, in conjunction with earlier findings, demonstrate that phycobiliprotein bilin lyases show high selectivity (rather than absolute specificity) for both the bilin and the polypeptide substrate. The rod substructures of the Anabaena sp. PCC 7120 phycobilisome contain the light harvesting proteins C-phycocyanin and phycoerythrocyanin (PEC). Even at low light intensities, PEC represents no more than 5% of the phycobilisome protein. The β subunits of both proteins carry thioether-linked phycocyanobilin (PCB) at β-Cys-82 and β-Cys-155; however, C-phycocyanin has PCB at α-Cys-84 whereas PEC α subunit carries phycobiliviolin at this position. The Anabaena sp. PCC 7120 pec operon is made up of five genes. PecB and pecA encode the β and α subunits of PEC, pecC encodes a linker polypeptide associated with PEC in the rod substructure, and pecE and pecF are genes of unknown function that show a high degree of homology to cpcE and cpcF, that encode a C-phycocyanin α subunit PCB lyase (Fairchild, C. D., Zhao, J., Zhou, J., Colson, S. E., Bryant, D. A., and Glazer, A. N.(1992) Proc. Natl. Acad. Sci. U. S. A. 89, 7017-7021). Insertional mutants in pecE and pecF, and an interposon mutant in which a portion of both pecE and pecF was deleted, were constructed. All three types of mutants grew 1.3 times slower than wild-type under limiting light conditions and showed a 20% reduction in the PCB content of whole cells relative to chlorophyll a. Holo-PEC was missing from the phycobilisomes of all three types of mutants and the level of the PEC linker polypeptide was reduced relative to the wild-type. However, ∼30% of the wild-type level of the PEC β subunit was present in all of these phycobilisomes. In contrast, the PEC α subunit was barely detectable in the pecE and pecF mutants, but was present in the pecEF deletion mutant as a PCB-adduct in a 1:1 ratio with the PEC β subunit. The identity of this “unnatural” adduct was confirmed by isolation of the subunit and amino-terminal sequencing. These biochemical results support the inference that pecE and pecF encode a PEC α subunit phycobiliviolin lyase, and, in conjunction with earlier findings, demonstrate that phycobiliprotein bilin lyases show high selectivity (rather than absolute specificity) for both the bilin and the polypeptide substrate.