Characterization of Pheophytin Ground States in Rhodobacter sphaeroides R26 Photosynthetic Reaction Centers from Multispin Pheophytin Enrichment and 2-D 13C MAS NMR Dipolar Correlati

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• 作者：Tatjana A. Egorova-Zachernyuk ; Barth van Rossum ; Gert-Jan Boender ; Eric Franken ; Jennifer Ashurst ; Jan Raap ; Peter Gast ; Arnold J. Hoff ; Hartmut Oschkinat ; and Huub J. M. de Groot
• 刊名：Biochemistry
• 出版年：1997
• 出版时间：June 17, 1997
• 年：1997
• 卷：36
• 期：24
• 页码：7513 - 7519
• 全文大小：252K
• 年卷期：v.36,no.24(June 17, 1997)
• ISSN：1520-4995

文摘

The electronic ground states of pheophytin cofactors potentiallyinvolved in symmetry breakingbetween the A and B branch for electron transport in the bacterialphotosynthetic reaction center havebeen investigated through a characterization of the electron densitiesat individual atomic positions ofpheophytin a from ¹³C chemical shift data.A new experimental approach involving multispin ¹³Clabelingand 2-D NMR is presented. Bacterial photosynthetic reactioncenters of Rhodobacter sphaeroides R26were reconstituted with uniformly ¹³C biosyntheticallylabeled (plant) Pheo a in the two pheophytinbindingsites. From the multispin labeled samples 1-D and 2-D solid-state¹³C magic angle spinning NMR spectracould be obtained and used to characterize the pheophytin aground state in the Rb. sphaeroides R26RCs, i.e., without a necessity for time-consuming selective labelingstrategies involving organic synthesis.From the 2-D solid state ¹³C-¹³Ccorrelation spectra collected with spinning speeds of 8 and 10kHz,with mixing times of 1 and 0.8 ms, many ¹³C resonances ofthe [U-¹³C]Pheo a moleculesreconstitutedin the RCs could be assigned in a single set of experiments. Partsof the pheophytins interacting with theprotein, at the level of ¹³C shifts modified by binding,could be identified. Small reconstitution shifts aredetected for the 17² side chain of ring IV. Incontrast, there is no evidence for electrostaticdifferencesbetween the two Pheo a, for instance, due to a possiblystrong selective electrostatic interaction with GluL104 on the active branch. The protonation states appear the same,and the NMR suggests a strongoverall similarity between the ground states of the two Pheoa, which is of interest in view of theasymmetryof the electron transfer.