Molecular cloning, characterization, and expression of a redox-responsive cutinase from Monilinia fructicola (Wint.) Honey

Fungal Genetics and Biology : FG & B
G Y WangRichard M Bostock

Abstract

cDNA clones encoding a cutinase expressed in cutin-induced cultures of the plant pathogen Monilinia fructicola were isolated using a protein-based strategy. The largest cDNA (Mfcut1) was found to contain an open reading frame of 603 bp that predicted a 20.2-kDa protein of 201 amino acids with a 20-amino-acid secretory signal peptide and a pI of 8.4. The predicted protein contained cutinase/lipase consensus sequences with active site serines and potential protein kinase phosphorylation sites. Comparison of the deduced amino sequence from Mfcut1 with other fungal cutinase sequences revealed new features, which include conserved cysteines, C-terminal aromatic residues, and a novel histidine substitution in the D-H active site motif. The presence in the growth medium of antioxidants, such as caffeic acid, suppressed mRNA accumulation and enzyme activity of a cutinase from M. fructicola. MFCUT1 was expressed at high levels as a His-tagged fusion protein in Pichia pastoris and purified to apparent homogeneity in a single step by Ni(2+)-nitrilotriacetic acid affinity chromatography. Analysis of variant MFCUT1 mutants in which the novel serine and histidine residues were replaced by site-directed mutagenesis indicated that these residu...Continue Reading

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Citations

Mar 23, 2010·Applied Microbiology and Biotechnology·Katja KoschorreckBernhard Hauer
Dec 29, 2012·Journal of Industrial Microbiology & Biotechnology·Shaoqing YangZhengqiang Jiang
May 17, 2007·Biotechnology Letters·Jacob Dam SørensenMarilyn G Wiebe
Jan 13, 2012·Applied Biochemistry and Biotechnology·Denise Castro-OchoaAmelia Farrés
Jul 29, 2008·The Journal of Biological Chemistry·Sheng ChenJian Chen
Jul 19, 2013·The Journal of General and Applied Microbiology·Kasturi DuttaVeeranki Venkata Dasu
Oct 15, 2013·Applied Microbiology and Biotechnology·Antti NyyssöläTiina Nakari-Setälä
Sep 24, 2013·Biotechnology Advances·Sheng ChenJing Wu
May 3, 2016·Journal of Agricultural and Food Chemistry·Leandro Oliveira LinoBénédicte Quilot-Turion
Sep 1, 2017·Preparative Biochemistry & Biotechnology·Ali Osman Adıgüzel, Münir Tunçer
Sep 5, 2019·Phytopathology·Ting-Hang LiuRichard M Bostock
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Jun 3, 2020·Molecular Plant-microbe Interactions : MPMI·Silvia RodríguezEduardo A Espeso
Sep 23, 2020·Food Science and Technology International = Ciencia Y Tecnología De Los Alimentos Internacional·Núria Baró-MontelJosep Usall

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