A full-length cDNA sequence of plant type CRY (designated Hae-P-CRY) was cloned from the green alga Haematococcus pluvialis. The cDNA sequence was 3608 base pairs (bp) in length, which contained a 2988-bp open reading frame encoding 995 amino acids with molecular mass of 107.7 kDa and isoelectric point of 6.19. Multiple alignment analysis revealed that the deduced amino acid sequence of Hae-P-CRY shared high identity of 47-66% with corresponding plant type CRYs from other eukaryotes. The catalytic motifs of plant type CRYs were detected in the amino acid sequence of Hae-P-CRY including the typical PHR and CTE domains. Phylogenetic analysis showed that the Hae-P-CRY was grouped together with other plant type CRYs from green algae and higher plants, which distinguished from other distinct groups. The transcriptional level of Hae-P-CRY was strongly decreased after 0-4 h under HL stress. In addition, the Hae-P-CRY gene was heterologously expressed in Escherichia coli BL21 (DE3) and successfully purified. The typical spectroscopic characteristics of plant type CRYs were present in Hae-P-CRY indicated that it may be an active enzyme, which provided valuable clue for further functional investigation in the green alga H. pluvialis. These results lay the foundation for further function and interaction protein identification involved in CRYs mediated signal pathway under HL stress in H. pluvialis.

中文翻译：

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绿藻血球菌的新型植物型隐花色素基因的克隆，表达和表征。

从绿藻类嗜血红球菌克隆了植物类型CRY的全长cDNA序列（命名为Hae-P-CRY）。cDNA序列长度为3608个碱基对（bp），其包含2988bp的开放阅读框，其编码995个氨基酸，分子量为107.7kDa，等电点为6.19。多重比对分析显示，推导的Hae-P-CRY氨基酸序列与其他真核生物的相应植物类型CRY具有47-6％的高度同一性。在Hae-P-CRY的氨基酸序列（包括典型的PHR和CTE域）中检测到了植物型CRY的催化基序。系统发育分析表明，Hae-P-CRY与来自绿藻和高等植物的其他植物类型的CRYs分组在一起，这与其他不同的群体有所区别。在HL胁迫下0-4小时后，Hae-P-CRY的转录水平强烈降低。此外，Hae-P-CRY基因在大肠杆菌BL21（DE3）中异源表达并成功纯化。Hae-P-CRY中存在植物型CRYs的典型光谱特征，表明它可能是一种活性酶，为绿藻H. pluvialis中的进一步功能研究提供了有价值的线索。这些结果为进一步的功能和相互作用蛋白鉴定参与了在幽门螺杆菌HL胁迫下CRYs介导的信号通路奠定了基础。Hae-P-CRY中存在植物型CRYs的典型光谱特征，表明它可能是一种活性酶，为绿藻H. pluvialis中的进一步功能研究提供了有价值的线索。这些结果为进一步的功能和相互作用蛋白鉴定参与了在幽门螺杆菌HL胁迫下CRYs介导的信号通路奠定了基础。Hae-P-CRY中存在植物型CRYs的典型光谱特征，表明它可能是一种活性酶，为绿藻H. pluvialis中的进一步功能研究提供了有价值的线索。这些结果为进一步的功能和相互作用蛋白鉴定参与了在幽门螺杆菌HL胁迫下CRYs介导的信号通路奠定了基础。