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首页> 外文期刊>Journal of bacteriology >Probing the CD Lumenal Loop Region of the D2 Protein of Photosystem II in Synechocystis sp. Strain PCC 6803 by Combinatorial Mutagenesis
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Probing the CD Lumenal Loop Region of the D2 Protein of Photosystem II in Synechocystis sp. Strain PCC 6803 by Combinatorial Mutagenesis

机译：探查集胞藻sp.Photosystem II D2蛋白的CD管腔环区。组合诱变株PCC 6803

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The CD lumenal loop region of the photosystem II reaction center protein D2 contains residues involved in oxygen evolution. Since detailed structural information about this region is unavailable, an M13-based combinatorial mutagenesis approach was used to investigate structure-function relationships in this vital region of D2 inSynechocystis sp. strain PCC 6803. The CD loop coding region contains close to 100 nucleotides, and for effective mutagenesis, it was subdivided into four regions of seven to eight codons. A gain-of-function selection protocol was employed such that all mutants that were selected contained a functional D2 protein. In this way, conservation patterns of residues along with numbers and types of amino acid substitutions accommodated at each position for each set of mutants would indicate which residues in the CD loop may play important structural and functional roles. Results of this study have substantiated the importance of residues previously studied by site-directed mutagenesis such as Arg180 and His189 and have identified other previously unremarkable residues in the CD loop (such as Ser166, Phe169, and Ala170) that cannot be replaced by many other residues. In addition, the pliability of the CD loop was further tested using deletion and D1-D2 substitution constructs in M13. This showed that the length of the loop was important to its function, and in two cases, D2 could accommodate homologous sequences from D1, which forms a heterodimer with D2 in photosystem II, but not the other way around. This study of the CD loop in D2 provides valuable clues regarding the structural and functional requirements of the region.
机译：光系统II反应中心蛋白D2的CD管腔环区域包含与氧释放有关的残基。由于无法获得有关该区域的详细结构信息，因此使用基于M13的组合诱变方法来研究Synechocystis sp中D2的这一重要区域的结构-功能关系。 CD环编码区包含近100个核苷酸，为了有效诱变，将其细分为7至8个密码子的四个区域。使用功能获得选择方案，以使所选择的所有突变体均含有功能性D2蛋白。以这种方式，残基的保守模式以及每组突变体在每个位置上容纳的氨基酸取代的数量和类型将指示CD环中的哪些残基可能起重要的结构和功能作用。这项研究的结果证实了先前通过定点诱变研究的残基（例如Arg180和His189）的重要性，并确定了CD环中其他先前不显着的残基（例如Ser166，Phe169和Ala170），这些残基无法被许多其他残基取代。残留物。另外，使用M13中的缺失和D1-D2取代构建体进一步测试了CD环的柔韧性。这表明环的长度对其功能很重要，在两种情况下，D2可以容纳来自D1的同源序列，后者在光系统II中与D2形成异二聚体，但反之则不然。对D2中CD回路的这项研究提供了有关该区域的结构和功能要求的宝贵线索。

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