Stewart D H, Cua A, Chisholm D A, Diner B A, Bocian D F, Brudvig G W
Department of Chemistry, Yale University, New Haven, Connecticut 06520-8107, USA.
Biochemistry. 1998 Jul 14;37(28):10040-6. doi: 10.1021/bi980668e.
Chlorophyll Z (ChlZ) is a redox-active chlorophyll (Chl) which is photooxidized by low-temperature (<100 K) illumination of photosystem II (PSII) to form a cation radical, ChlZ+. This cofactor has been proposed to be an "accessory" Chl in the PSII reaction center and is expected to be buried in the transmembrane region of the PSII complex, but the location of ChlZ is unknown. A series of single-replacement site-directed mutants of PSII were made in which each of two potentially Chl-ligating histidines, D1-H118 or D2-H117, was substituted with amino acids which varied in their ability to coordinate Chl. Assays of the wild-type and mutant strains showed parallel phenotypes for the D1-118 and D2-117 mutants: noncoordinating or poorly coordinating residues at either position decreased photosynthetic competence and impaired assembly of PSII complexes. Only the mutants substituted with glutamine (D1-H118Q and D2-H117Q) had phenotypes comparable to the wild-type strain. The ChlZ+ cation was characterized by low-temperature electron paramagnetic resonance (EPR), near-infrared (IR) absorbance, and resonance Raman (RR) spectroscopies in wild-type, H118Q, and H117Q PSII core complexes. The quantum yield of ChlZ+ formation is the same (approximately 2.5% per saturating flash at 77 K) for wild-type, H118Q, and H117Q, indicating that its efficiency of photooxidation is unchanged by the mutations. Similarly, the EPR and near-IR absorbance spectra of ChlZ+ are insensitive to the mutations made at D1-118 and D2-117. In contrast, the RR signature of ChlZ+ in H118Q PSII, obtained by selective near-IR excitation into the ChlZ+ cation absorbance band, is significantly altered relative to wild-type PSII while the RR spectrum of ChlZ+ in the H117Q mutant remains identical to wild-type. Shifts in the RR spectrum of ChlZ+ in H118Q reflect a change in the structure of the Chl ring, most likely due to a perturbation of the core size and/or extent of doming caused by a change in the axial ligand to Mg(II). Thus, we conclude that the axial ligand to ChlZ is H118 of the D1 polypeptide. Furthermore, we propose that H117 of the D2 polypeptide is the ligand to a homologous redox-inactive accessory Chl which we term ChlD. The Chl Z and D terminology reflects the 2-fold structural symmetry of PSII which is apparent in the redox-active tyrosines, YZ and YD, and the active/inactive branch homology of the D1/D2 polypeptides with the L/M polypeptides of the bacterial reaction center.
叶绿素Z（ChlZ）是一种具有氧化还原活性的叶绿素（Chl），通过光系统II（PSII）在低温（<100 K）光照下发生光氧化形成阳离子自由基ChlZ+。这种辅因子被认为是PSII反应中心的一种“辅助”叶绿素，预计埋在PSII复合物的跨膜区域，但ChlZ的位置尚不清楚。构建了一系列PSII的单取代定点突变体，其中两个潜在的叶绿素结合组氨酸D1-H118或D2-H117分别被配位能力不同的氨基酸取代。对野生型和突变菌株的分析表明，D1-118和D2-117突变体具有相似的表型：任一位置的非配位或配位能力差的残基都会降低光合能力并损害PSII复合物的组装。只有用谷氨酰胺取代的突变体（D1-H118Q和D2-H117Q）具有与野生型菌株相当的表型。通过低温电子顺磁共振（EPR）、近红外（IR）吸收和共振拉曼（RR）光谱对野生型、H118Q和H117Q PSII核心复合物中的ChlZ+阳离子进行了表征。野生型、H118Q和H117Q的ChlZ+形成量子产率相同（在77 K下每个饱和闪光约为2.5%），表明其光氧化效率不受突变影响。同样，ChlZ+的EPR和近红外吸收光谱对D1-118和D2-117处的突变不敏感。相比之下，通过选择性近红外激发进入ChlZ+阳离子吸收带获得的H118Q PSII中ChlZ+的RR特征相对于野生型PSII有显著变化，而H117Q突变体中ChlZ+的RR光谱与野生型保持相同。H118Q中ChlZ+的RR光谱变化反映了叶绿素环结构的变化，最可能是由于轴向配体与Mg(II)的变化导致核心大小和/或穹顶程度的扰动。因此，我们得出结论，ChlZ的轴向配体是D1多肽的H118。此外，我们提出D2多肽的H117是一种同源的氧化还原无活性辅助叶绿素（我们称之为ChlD）的配体。Chl Z和D的术语反映了PSII的2倍结构对称性，这在氧化还原活性酪氨酸YZ和YD以及D1/D2多肽与细菌反应中心的L/M多肽的活性/非活性分支同源性中很明显。
