Morita Tomotake, Konishi Masaaki, Fukuoka Tokuma, Imura Tomohiro, Kitamoto Dai
Research Institute for Innovations in Sustainable Chemistry, National institute of Advanced Industrial Science and Technology, AIST, Tsukuba Central 5-2, Higashi 1-1, Tsukuba, Ibaraki 305-8565, Japan.
Yeast. 2006 Jul 15;23(9):661-71. doi: 10.1002/yea.1386.
Pseudozyma antarctica T-34 secretes a large amount of biosurfactants (BS), mannosylerythritol lipids (MEL), from different carbon sources such as hydrocarbons and vegetable oils. The detailed biosynthetic pathway of MEL remained unknown due to lack of genetic information on the anamorphic basidiomycetous yeasts, including the genus Pseudozyma. Here, in order to obtain genetic information on P. antarctica T-34, we constructed a cDNA library from yeast cells producing MEL from soybean oil and identified the genes expressed through the creation of an expressed sequence tags (EST) library. We generated 398 ESTs, assembled into 146 contiguous sequences. Based upon a BLAST search similarity cut-off of E<or=10(-5), 21.4% of all contigs were orphan, while 78.6% showed similarity to sequences in the protein database; 60.3% of all contiguous sequences shared significant identities to hypothetical protein of Ustilago maydis, which is a smut fungus and BS producer. Based on the gene expression study using real-time reverse transcriptase-PCR, the predicted genes, such as mannosyltranferase and acyltransferase, were demonstrated to be highly involved in MEL biosynthesis in soybean oil-grown cells.
南极假丝酵母T-34能从烃类和植物油等不同碳源中分泌大量生物表面活性剂（BS），即甘露糖赤藓糖醇脂（MEL）。由于包括假丝酵母属在内的无性型担子菌酵母缺乏遗传信息，MEL的详细生物合成途径仍不清楚。在此，为了获得南极假丝酵母T-34的遗传信息，我们从以大豆油生产MEL的酵母细胞构建了一个cDNA文库，并通过创建一个表达序列标签（EST）文库来鉴定所表达的基因。我们生成了398个EST，组装成146个连续序列。基于E≤10^(-5)的BLAST搜索相似性截止值，所有重叠群中有21.4%是孤儿序列，而78.6%与蛋白质数据库中的序列显示出相似性；所有连续序列中有60.3%与黑粉菌和BS生产者玉米黑粉菌的假定蛋白具有显著的同源性。基于使用实时逆转录PCR的基因表达研究，预测的基因，如甘露糖基转移酶和酰基转移酶，被证明在以大豆油培养的细胞中高度参与MEL的生物合成。
