| Summary: | 博士 === 國立中興大學 === 生物科技學研究所 === 101 === Cytochrome (Cyt) b559 is one of the essential photosystem II (PSII) proteins and is composed of α and β heme-bridged heterodimer proteins (encoded by psb E and psb F genes, respectively). Previous studies have proposed that Cyt b559 participates in secondary electron transfers that protect PSII from photoinhibition by forming a cyclic electron-transfer pathway within PSII. However, the mechanism remains elusive. We identified a spontaneously-generated mutant from Synechocystis sp. PCC6803 wild-type cells grown in BG-11 agar plates containing 5 mM glucose and 10 μM DCMU. This mutant carries an R7L mutation on the α-subunit of cyt b559 in PSII. This mutant grew at about the same rate as wild-type cells under photoautotrophical conditions but grew faster than wild-type cells under photoheterotrophical conditions. 77 K fluorescence and 295 K chlorophyll a fluorescence spectral results indicated that the energy delivery from phycobilisomes to PSII reaction centers was partially inhibited or uncoupled in this mutant. We propose that the Arg7Leu mutation on the α-subunit of cyt b559 alters the interaction between the APC core complex and PSII reaction centers, which reduces energy delivery from the antenna to the reaction center and thus protects mutant cells from DCMU-induced photo-oxidative stress.
To provide the insight into the structural and functional roles of cyt b559, we created a series of site-directed mutants on the charged residues (Glu6, Arg7, Asp11 and Arg 17 of the α subunit and Arg17 of the β subunit) on the cytoplasmic side of cyt b559. All of the mutant cells grew photoautotrophically and assembled stable photosystem II. However, R7Eα, R17Eα and R17Lβ mutant cells grew significantly slower and were more susceptible to photoinhibition than wild-type cells. Our results demonstrate that the electrostatic interactions between these arginine residues and the heme propionates of cytochrome b559 are important to the structure and redox properties of cytochrome b559. In addition, the interaction of the allophycocyanin core complex with the orange carotenoid protein was altered in Arg mutant cells. Furthermore, UPLC-APCI-QTOFMS results showed that the PQ pool was more reduced in R7Eα and R17Lβ mutant cells than wild-type cells in the dark. The implications of our findings on the structure and physiological functions of Cyt b559 in PSII will be discussed.
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