Nguyen Long Nam, Dao Tuyet Thi, Živković Tijana, Fehrholz Markus, Schäfer Wilhelm, Salomon Siegfried
University of Hamburg, Biocenter Klein Flottbek, Department of Molecular Phytopathology and Genetics, Ohnhorststr. 18, D-22609 Hamburg, Germany; Vietnam Academy of Science and Technology, Institute of Biotechnology, 10600 Hanoi, Viet Nam.
Vietnam Academy of Science and Technology, Institute of Biotechnology, 10600 Hanoi, Viet Nam.
Enzyme Microb Technol. 2010 May 5;46(6):479-86. doi: 10.1016/j.enzmictec.2010.02.005. Epub 2010 Feb 20.
Fusarium graminearum is a filamentous fungus that causes devastating diseases on plants of economic importance including maize, wheat, and barley. F. graminearum is able to utilize triglycerides as a carbon source during growth. Extracellular lipases are the preferred enzymes to catalyze the hydrolysis of fats and oils. Lipases are ubiquitous enzymes of considerable physiological significance and industrial potential. Previously, FGL1 was the first described F. graminearum extracellular lipase associated with virulence. We report the biochemical characterization of FGL1 and four new secreted lipases of F. graminearum. The lipase genes of F. graminearum wild-type strain 8/1 were sequenced, cloned and over-expressed in Pichia pastoris. We show that the lipases have their temperature optimum between 30 and 40°C and a pH optimum of ∼7. A broad range of lipase substrates, from C4 to C18 p-nitrophenyl esters, were hydrolyzed efficiently by the lipases, indicating the true lipolytic activity of the enzymes. Expression patterns of these lipases were also analyzed by semiquantitative RT-PCR in F. graminearum cultured in water supplemented with 2% (v/v) wheat germ oil at 28°C. Transcripts of all examined lipases are detectable and the genes are regulated differently under these culture conditions. Our data indicated that F. graminearum possesses a ubiquitous source of secreted lipases, which could be used for industrial intentions. We also provided the foundation of lipase expression in vitro, which is necessary for further characterization.
禾谷镰刀菌是一种丝状真菌，可对包括玉米、小麦和大麦在内的具有经济重要性的植物引发毁灭性病害。禾谷镰刀菌在生长过程中能够利用甘油三酯作为碳源。细胞外脂肪酶是催化油脂水解的首选酶。脂肪酶是一类普遍存在的酶，具有相当重要的生理意义和工业潜力。此前，FGL1是首次被描述的与禾谷镰刀菌毒力相关的细胞外脂肪酶。我们报道了FGL1以及禾谷镰刀菌四种新分泌脂肪酶的生化特性。对禾谷镰刀菌野生型菌株8/1的脂肪酶基因进行了测序、克隆，并在毕赤酵母中进行了过表达。我们发现这些脂肪酶的最适温度在30至40°C之间，最适pH约为7。这些脂肪酶能有效水解从C4到C18的对硝基苯酯等多种脂肪酶底物，表明这些酶具有真正的脂解活性。还通过半定量RT-PCR分析了这些脂肪酶在28°C下于添加2%（v/v）小麦胚芽油的水中培养的禾谷镰刀菌中的表达模式。所有检测的脂肪酶转录本均可检测到，并且在这些培养条件下基因的调控方式不同。我们的数据表明，禾谷镰刀菌拥有丰富的分泌脂肪酶来源，可用于工业用途。我们还为脂肪酶的体外表达奠定了基础，这对于进一步的特性研究是必要的。
