深海沉积物宏基因组中新型对羟基苯丙酮酸双加氧酶基因的鉴定

Identification of novel 4-hydroxyphenylpyruvate dioxygenase genes from the deep sea sediment metagenomes

摘要: 为采用不依赖培养的方法获取深海沉积物微生物遗传资源，利用深海沉积物DNA构建了2个宏基因组文库，对对羟基苯丙酮酸双加氧酶（4HPPD）活性进行了筛选，并分离出3个克隆子。体外转座子突变和全长测序发现了两个新的4HPPD基因和一个新的尿黑酸（HGA）代谢基因簇。克隆到 pMD18-T 载体中的3个新的4HPPD 基因能够在大肠杆菌中产生水溶性棕色素。该色素被鉴定为黑色素，并与 HGA 相关。结果表明，深海沉积物宏基因组含有多样性丰富的4HPPD功能基因，可用于发现新的黑色素生物合成基因。

Abstract: To access the microbial genetic resources of deep-sea sediments by a culture-independent approach, two metagenomic libraries constructed with deep sea sediments DNA were screened for the 4-hydroxyphenylpyruvate dioxygenase (4HPPD) activity and three positive clones were obtained. In vitro transposon mutagenesis and full-length sequencing revealed two novel 4HPPD genes and a novel homogentistic acid (HGA) metabolism gene cluster. Three new4HPPD genes cloned into pMD18-T vector could produce water-soluble brown pigment in Escherichia coli. The pigment was identified to be pyomelanin and associated with HGA. Results show that the metagenomes of deep-sea sediments contain diverse4HPPD functional genes, which can be used to find new melanin biosysthesis genes.