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首页> 外文期刊>Journal of Experimental Botany >Cloning, functional expression, and characterization of a chalcone 3-hydroxylase from Cosmos sulphureus
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Cloning, functional expression, and characterization of a chalcone 3-hydroxylase from Cosmos sulphureus

机译：硫磺波斯菊查尔酮3-羟化酶的克隆，功能表达和表征

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摘要

A chalcone 3-hydroxylase (CH3H) cDNA clone was isolated and characterized from Cosmos sulphureus petals accumulating butein (2', 3,4,4'-tetrahydroxychalcone) derivatives as yellow flower pigments. The recombinant protein catalyses the introduction of an additional hydroxyl group in the B-ring of chalcones, a reaction with high similarity to the hydroxylation of flavonoids catalysed by the well-studied flavonoid 3'-hydroxylase (F3'H). CH3H shows high specificity for chalcones, but a low F3'H activity was also detected. By contrast, the common F3'H from C. sulphureus does not accept chalcones as substrates and is therefore unlikely to be involved in the creation of the B-ring hydroxylation pattern of the yellow flower pigments. CH3H was primarily expressed in young buds, the main tissue for chalcone pigment formation. Expression levels in open flowers and 3-d-old seedlings were lower and almost no CH3H expression was observed in leaves. F3'H, in contrast, showed the highest expression also in buds, but comparable expression rates in all other tissues tested. Recombinant hybrid proteins constructed from CH3H and F3'H fragments demonstrated that amino acid residues at a substrate recognition site and an insertion of four amino acid residues in a putative loop region have an impact on chalcone acceptance. This is the first identification of a CH3H cDNA from any plant species.
机译：从积累了丁酮（2'，3,4,4'-四羟基查尔酮）衍生物的黄色花色素的硫磺波斯菊花瓣中分离并鉴定了查尔酮3-羟化酶（CH3H）cDNA克隆。重组蛋白催化在查耳酮的B环中引入额外的羟基，该反应与经过充分研究的类黄酮3'-羟化酶（F3'H）催化的类黄酮的羟基化反应高度相似。 CH3H对查耳酮显示出高特异性，但也检测到F3'H活性低。相比之下，来自硫脲梭菌的常见F3'H不接受查耳酮作为底物，因此不太可能参与黄色花色素的B环羟基化图案的产生。 CH3H主要在幼芽中表达，幼芽是查尔酮色素形成的主要组织。在开放花和3 d幼苗中的表达水平较低，并且在叶片中几乎没有观察到CH3H表达。相比之下，F3'H在芽中也显示最高的表达，但是在所有其他测试组织中的表达率相当。由CH3H和F3'H片段构建的重组杂合蛋白表明，底物识别位点的氨基酸残基以及推定的环区域中四个氨基酸残基的插入对查耳酮的接受有影响。这是来自任何植物物种的CH3H cDNA的首次鉴定。

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