In vivo system for analyzing the function of the PsbP protein using Chlamydomonas reinhardtii.
The PsbP protein is an extrinsic subunit of photosystem II (PSII) specifically developed in green-plant species including land plants and green algae. The protein-protein interactions involving PsbP and its effect on oxygen evolution have been investigated in vitro using isolated PSII membranes. However, the importance of those interactions needs to be examined at the cellular level. To this end, we developed a system expressing exogenous PsbP in the background of the Chlamydomonas BF25 mutant lacking native PsbP. Expression of His-tagged PsbP successfully restored the oxygen-evolving activity and photoautotrophic growth of the mutant, while PsbP-∆15 lacking the N-terminal 15 residues, which are crucial for the oxygen-evolving activity of spinach PSII in vitro, only partially did. This demonstrated the importance of N-terminal sequence of PsbP for the photosynthetic activity in vivo. Furthermore, the PSII-LHCII supercomplex can be specifically purified from the Chlamydomonas cells having His-tagged PsbP using a metal affinity chromatography. This study provides a platform not only for the functional analysis of PsbP in vivo but also for structural analysis of the PSII-LHCII supercomplex from green algae.
Graduate School of Biostudies, Kyoto University, Sakyo, Kyoto, 606-8502, Japan.
Taishi
http://orcid.org/0000-0003-2831-7010
T
Nishimura
Graduate School of Biostudies, Kyoto University, Sakyo, Kyoto, 606-8502, Japan.
Fumihiko
F
Sato
Graduate School of Biostudies, Kyoto University, Sakyo, Kyoto, 606-8502, Japan. ifuku@kais.kyoto-u.ac.jp.
Kentaro
http://orcid.org/0000-0003-0241-8008
K
Ifuku
D001483:Base Sequence; D016825:Chlamydomonas reinhardtii / Q000235:genetics / Q000378:metabolism*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D005816:Genetic Complementation Test; D006639:Histidine / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D063347:Native Polyacrylamide Gel Electrophoresis; D009842:Oligopeptides / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D045332:Photosystem II Protein Complex / Q000235:genetics / Q000378:metabolism*; D020524:Thylakoids / Q000378:metabolism
C471213:His-His-His-His-His-His; D009842:Oligopeptides; D010455:Peptides; D045332:Photosystem II Protein Complex; D006639:Histidine; D010100:Oxygen
Chlamydomonas reinhardtii; His-tag; Photosystem II; PsbP