A novel recombinant chlorophyllase from cyanobacterium Cyanothece sp. ATCC 51142 for the production of bacteriochlorophyllide a
A novel recombinant chlorophyllase from cyanobacterium Cyanothece sp. ATCC 51142 for the production of bacteriochlorophyllide a
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DOI：
10.1002/bab.1380
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发表时间：
2016-05-01
影响因子：
2.8
通讯作者：
Shaw, Jei-Fu
中科院分区：
文献类型：
--
作者：
Chou, Yi-Li;Lee, Ya-Lin;Shaw, Jei-Fu
芽孢杆菌脱镁叶绿体a(BPheid A)是细菌素a(BCA)的前体，可用于体外和体内生物化学的光动力治疗。本研究成功地分离和表达了一株光合作用细菌(Cyanothess sp.ATCC 51142)叶绿素酶，称为氰基CLH，可用作生物催化剂，通过降解细菌叶绿素a(BCHLA)来生产苯丙氨酸前体。底物专一性和酶动力学分析结果表明，重组氰基CLH更倾向于水解Bchl a生成细菌叶绿素a(BChlide A)，通过去除镁离子可将其转化为BPheid a。克隆并在大肠杆菌BL-21(DE3)中表达，其相对分子质量为54.7 kDa。其推导的氨基酸序列包含一个独特的脂肪酶基序GHSLG，它不同于其他植物的GHSRG序列，并且缺乏典型和保守的催化三联体Ser-Asp-His的组氨酸。对重组氰基CLH进行了生化分析，结果表明其最适pH为7.0，最适温度为60℃。
Bacteriopheophorbide a (BPheid a) is used as a precursor for bacteriochlorin a (BCA), which can be used for photodynamic therapy in both in vitro and in vivo biochemical applications. This study successfully isolated and expressed a photosynthetic bacterium (Cyanothece sp. ATCC 51142) chlorophyllase called CyanoCLH, which can be used as a biocatalyst for the production of a BCA precursor by degrading bacteriochlorophyll a (BChl a). Substrate specificity and enzyme kinetic analyses were performed and the results verified that the recombinant CyanoCLH preferred hydrolyzing BChl a to produce bacteriochlorophyllide a (BChlide a), which can be converted to BPheid a by removing magnesium ion. The recombinant CyanoCLH was cloned and expressed in Escherichia coli BL-21 (DE3), and its molecular weight was 54.7kDa. The deduced amino acid sequence of the recombinant CyanoCLH comprised a unique lipase-motif GHSLG, which differs from the GHSRG sequence of other plants and lacks a histidine of the typical and conserved catalytic triad Ser-Asp-His. The recombinant CyanoCLH was subjected to biochemical analyses, and the results indicated that its optimal pH and temperature were 7.0 and 60 degrees C, respectively.