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The D1-V185N mutation alters substrate water exchange by stabilizing alternative structures of the Mn4Ca-cluster in photosystem II
Biochimica et Biophysica Acta (BBA) - Bioenergetics ( IF 3.4 ) Pub Date : 2020-09-23 , DOI: 10.1016/j.bbabio.2020.148319
Casper de Lichtenberg , Anton P. Avramov , Minquan Zhang , Fikret Mamedov , Robert L. Burnap , Johannes Messinger

In photosynthesis, the oxygen-evolving complex (OEC) of the pigment-protein complex photosystem II (PSII) orchestrates the oxidation of water. Introduction of the V185N mutation into the D1 protein was previously reported to drastically slow O2-release and strongly perturb the water network surrounding the Mn4Ca cluster. Employing time-resolved membrane inlet mass spectrometry, we measured here the H218O/H216O-exchange kinetics of the fast (Wf) and slow (Ws) exchanging substrate waters bound in the S1, S2 and S3 states to the Mn4Ca cluster of PSII core complexes isolated from wild type and D1-V185N strains of Synechocystis sp. PCC 6803. We found that the rate of exchange for Ws was increased in the S1 and S2 states, while both Wf and Ws exchange rates were decreased in the S3 state. Additionally, we used EPR spectroscopy to characterize the Mn4Ca cluster and its interaction with the redox active D1-Tyr161 (YZ). In the S2 state, we observed a greatly diminished multiline signal in the V185N-PSII that could be recovered by addition of ammonia. The split signal in the S1 state was not affected, while the split signal in the S3 state was absent in the D1-V185N mutant. These findings are rationalized by the proposal that the N185 residue stabilizes the binding of an additional water-derived ligand at the Mn1 site of the Mn4Ca cluster via hydrogen bonding. Implications for the sites of substrate water binding are discussed.



中文翻译:

D1-V185N突变通过稳定光系统II中Mn 4 Ca簇的替代结构来改变底物水交换。

在光合作用中,色素-蛋白质复合物光系统II(PSII)的析氧复合物(OEC)协调水的氧化。以前有报道称将V185N突变引入D1蛋白会极大地减慢O 2释放并强烈干扰Mn 4 Ca团簇周围的水网络。利用时间分辨膜入口质谱法,我们在这里测量了结合在S 1,S 2和S上的快速(W f)和慢速(W s)交换底物水的H 2 18 O / H 2 16 O交换动力学。S 3表示Mn 4Ca簇的PSII核心复合物分离自野生型和集藻的D1-V185N菌株。PCC 6803。我们发现,W s的汇率在S 1和S 2状态下增加,而W f和W s汇率在S 3状态下都下降。此外,我们使用EPR光谱来表征Mn 4 Ca团簇及其与氧化还原活性D1-Tyr161(Y Z)的相互作用。在S 2状态下,我们观察到V185N-PSII中的多行信号大大减小,可以通过添加氨来恢复该信号。S中的分割信号D1-V185N突变体不影响1状态,而S 3状态的分裂信号不存在。这些发现通过N185残基通过氢键稳定了Mn 4 Ca簇的Mn1位点处额外的水衍生配体的结合的提议而变得合理。讨论了底物水结合位点的含义。

更新日期:2020-10-07
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