The Study On Enzymes Engineering Of Phycobiliproteins

In cyanobacteria and red algae, the main accessory lightharvesting complexes are the phycobilisomes (PBS), which attached to the cytoplasmic surface of the thylakoid membrane. These supramolecular protein complexes are primarily composed of phycobiliproteins, a brilliantly colored family of proteins bearing covalently attached, open-chain tetrapyrroles known as phycobilins. In additional, phycobilisomes also contain smaller amounts of linker peptides, which are required for proper assembly and functional organization of phycobilisomes. Phycobilin chromophores are generally bound to apoprotein at conserved positions by cysteinyl thioether linkages to generate phycobiliproteins.There are four cysteine in theα-phycoerythrin of Synechococcus sp. WH 8102. The seventy-fifth cysteine could covalently bound to the phycocyanobilin(PCB) via relatively stable thioether bonds. In vivo, the correct attachment of most chromophores is catalyzed by binding-site and chromophore-specific lyases. Recently,the RpcG encoded by gene synw2025 was proved to be a lyase, which could not only attach PEB to RpcA but also simultaneously isomerize it to PUB. Based on homology analysis, CpeZ encoded by gene synw2012 and CpeY encoded by gene synw2013 are highly similar to RpcG. Successfully the author cloned the cpeY and cpeZ to express vectors, which can express their own proteins in order to find out the new lyase.Gene coding for the lysase, CpcS, from Gloeobacter violaceus PCC 7421, was overexpressed in Escherichia coli. CpcS acts as a phycobiliprotein lyase that can attach, in vivo, phycocyanobilin (PCB) to the apo-β-subunits of C-phycocyanin (CpcB).The effect of nitrogen (N) starvation leads to a dramatic change in cell color from the normal blue-green to yellow-green, which is known as bleaching or chlorosis, in Synechocystis sp. PCC 6803. Pycobilisome(PBS) is progressively, rapidly, and almost completely degraded. In this sense, PC, the major constituant of PBS, acts as a nitrogen store. The transcripion of the genes were tested by quantitative real-time PCR following reverse transcription of total RNA. We confirmed that during the early stage of N deprivation, the abundance of clpP transcripts was significantly increased, while that of hhoB was significantly increased in the 20hrs, which the PBS was almost completely degraded.