Bode R

References (4)

Title : Three New Cutinases from the Yeast Arxula adeninivorans That Are Suitable for Biotechnological Applications - Bischoff_2015_Appl.Environ.Microbiol_81_5497
Author(s) : Bischoff F , Litwinska K , Cordes A , Baronian K , Bode R , Schauer F , Kunze G
Ref : Applied Environmental Microbiology , 81 :5497 , 2015
Abstract : The genes ACUT1, ACUT2, and ACUT3, encoding cutinases, were selected from the genomic DNA of Arxula adeninivorans LS3. The alignment of the amino acid sequences of these cutinases with those of other cutinases or cutinase-like enzymes from different fungi showed that they all had a catalytic S-D-H triad with a conserved G-Y-S-Q-G domain. All three genes were overexpressed in A. adeninivorans using the strong constitutive TEF1 promoter. Recombinant 6x His (6h)-tagged cutinase 1 protein (p) from A. adeninivorans LS3 (Acut1-6hp), Acut2-6hp, and Acut3-6hp were produced and purified by immobilized-metal ion affinity chromatography and biochemically characterized using p-nitrophenyl butyrate as the substrate for standard activity tests. All three enzymes from A. adeninivorans were active from pH 4.5 to 6.5 and from 20 to 30 degrees C. They were shown to be unstable under optimal reaction conditions but could be stabilized using organic solvents, such as polyethylene glycol 200 (PEG 200), isopropanol, ethanol, or acetone. PEG 200 (50%, vol/vol) was found to be the best stabilizing agent for all of the cutinases, and acetone greatly increased the half-life and enzyme activity (up to 300% for Acut3-6hp). The substrate spectra for Acut1-6hp, Acut2-6hp, and Acut3-6hp were quite similar, with the highest activity being for short-chain fatty acid esters of p-nitrophenol and glycerol. Additionally, they were found to have polycaprolactone degradation activity and cutinolytic activity against cutin from apple peel. The activity was compared with that of the 6x His-tagged cutinase from Fusarium solani f. sp. pisi (FsCut-6hp), also expressed in A. adeninivorans, as a positive control. A fed-batch cultivation of the best Acut2-6hp-producing strain, A. adeninivorans G1212/YRC102-ACUT2-6H, was performed and showed that very high activities of 1,064 U ml(-1) could be achieved even with a nonoptimized cultivation procedure.
ESTHER : Bischoff_2015_Appl.Environ.Microbiol_81_5497
PubMedSearch : Bischoff_2015_Appl.Environ.Microbiol_81_5497
PubMedID: 26048925
Gene_locus related to this paper: blaad-a0a060t5x1 , blaad-a0a060t512 , blaad-a0a060t6b2

Title : Secreted fungal effector lipase releases free fatty acids to inhibit innate immunity-related callose formation during wheat head infection - Blumke_2014_Plant.Physiol_165_346
Author(s) : Blumke A , Falter C , Herrfurth C , Sode B , Bode R , Schafer W , Feussner I , Voigt CA
Ref : Plant Physiol , 165 :346 , 2014
Abstract : The deposition of the (1,3)-beta-glucan cell wall polymer callose at sites of attempted penetration is a common plant defense response to intruding pathogens and part of the plant's innate immunity. Infection of the Fusarium graminearum disruption mutant Deltafgl1, which lacks the effector lipase FGL1, is restricted to inoculated wheat (Triticum aestivum) spikelets, whereas the wild-type strain colonized the whole wheat spike. Our studies here were aimed at analyzing the role of FGL1 in establishing full F. graminearum virulence. Confocal laser-scanning microscopy revealed that the Deltafgl1 mutant strongly induced the deposition of spot-like callose patches in vascular bundles of directly inoculated spikelets, while these callose deposits were not observed in infections by the wild type. Elevated concentrations of the polyunsaturated free fatty acids (FFAs) linoleic and alpha-linolenic acid, which we detected in F. graminearum wild type-infected wheat spike tissue compared with Deltafgl1-infected tissue, provided clear evidence for a suggested function of FGL1 in suppressing callose biosynthesis. These FFAs not only inhibited plant callose biosynthesis in vitro and in planta but also partially restored virulence to the Deltafgl1 mutant when applied during infection of wheat spikelets. Additional FFA analysis confirmed that the purified effector lipase FGL1 was sufficient to release linoleic and alpha-linolenic acids from wheat spike tissue. We concluded that these two FFAs have a major function in the suppression of the innate immunity-related callose biosynthesis and, hence, the progress of F. graminearum wheat infection.
ESTHER : Blumke_2014_Plant.Physiol_165_346
PubMedSearch : Blumke_2014_Plant.Physiol_165_346
PubMedID: 24686113

Title : Atan1p-an extracellular tannase from the dimorphic yeast Arxula adeninivorans: molecular cloning of the ATAN1 gene and characterization of the recombinant enzyme - Boer_2009_Yeast_26_323
Author(s) : Boer E , Bode R , Mock HP , Piontek M , Kunze G
Ref : Yeast , 26 :323 , 2009
Abstract : The tannase-encoding Arxula adeninivorans gene ATAN1 was isolated from genomic DNA by PCR, using as primers oligonucleotide sequences derived from peptides obtained after tryptic digestion of the purified tannase protein. The gene harbours an ORF of 1764 bp, encoding a 587-amino acid protein, preceded by an N-terminal secretion sequence comprising 28 residues. The deduced amino acid sequence was similar to those of tannases from Aspergillus oryzae (50% identity), A. niger (48%) and putative tannases from A. fumigatus (52%) and A. nidulans (50%). The sequence contains the consensus pentapeptide motif (-Gly-X-Ser-X-Gly-) which forms part of the catalytic centre of serine hydrolases. Expression of ATAN1 is regulated by the carbon source. Supplementation with tannic acid or gallic acid leads to induction of ATAN1, and accumulation of the native tannase enzyme in the medium. The enzymes recovered from both wild-type and recombinant strains were essentially indistinguishable. A molecular mass of approximately 320 kDa was determined, indicating that the native, glycosylated tannase consists of four identical subunits. The enzyme has a temperature optimum at 35-40 degrees C and a pH optimum at approximately 6.0. The enzyme is able to remove gallic acid from both condensed and hydrolysable tannins. The wild-type strain LS3 secreted amounts of tannase equivalent to 100 U/l under inducing conditions, while the transformant strain, which overexpresses the ATAN1 gene from the strong, constitutively active A. adeninivorans TEF1 promoter, produced levels of up to 400 U/l when grown in glucose medium in shake flasks.
ESTHER : Boer_2009_Yeast_26_323
PubMedSearch : Boer_2009_Yeast_26_323
PubMedID: 19387973
Gene_locus related to this paper: blaad-b7vfd0

Title : An extracellular lipase from the dimorphic yeast Arxula adeninivorans: molecular cloning of the ALIP1 gene and characterization of the purified recombinant enzyme - Boer_2005_Yeast_22_523
Author(s) : Boer E , Mock HP , Bode R , Gellissen G , Kunze G
Ref : Yeast , 22 :523 , 2005
Abstract : The lipase-encoding Arxula adeninivorans ALIP1 gene was isolated using fragments of lipase isolates obtained by trypsin digestion for the definition of oligonucleotide primers in a PCR screening approach. The gene harbours an ORF of 1347 bp encoding a 420 amino acid protein of some 50 kDa preceded by an N-terminal 28 prepro-secretion sequence. The deduced amino acid sequence was found to be similar to the lipases from Candida albicans and C. parapsilosis (34-38% identity) and more distantly related to other lipases. The sequence contains the consensus pentapeptide motif (-Gly-X-Ser-X-Gly-) that forms a part of the interfacial lipid recognition site in lipases. The expression of the gene is regulated by carbon source. In media supplemented with Tween 20, induction of the ALIP1 gene and accumulation of the encoded lipase in the medium is observed, thus demonstrating gene regulation by lipophilic compounds. The enzyme characteristics are analysed from isolates of native strains as well as from those of recombinant strains expressing the ALIP1 gene under control of the strong A. adeninivorans-derived TEF1 promoter. For both proteins a molecular mass of 100 kDa was determined, indicating a dimeric structure, a pH optimum at pH 7.5 and a temperature optimum at 30 degrees C. The enzyme hydrolyses all ester bonds in all triglyceride substrates tested. Middle-sized chain fatty acids are more efficiently hydrolysed than short- and long-chain fatty acids, with the highest activity on C8/C10 fatty acid esters pNP-caprylate, pNP-caprate and tricaprylin.
ESTHER : Boer_2005_Yeast_22_523
PubMedSearch : Boer_2005_Yeast_22_523
PubMedID: 15942926
Gene_locus related to this paper: arxad-q5gmi6