title: Genomic and secretomic insight into lignocellulolytic system of an endophytic bacterium Pantoea ananatis Sd-1.
abstract: BACKGROUND
  Exploring microorganisms especially bacteria associated with the degradation of lignocellulosic biomass shows great potentials in biofuels production. The rice endophytic bacterium Pantoea ananatis Sd-1 with strong lignocellulose degradation capacity has been reported in our previous study. However, a comprehensive analysis of its corresponding degradative system has not yet been conducted. The aim of this work is to identify and characterize the lignocellulolytic enzymes of the bacterium to understand its mechanism of lignocellulose degradation and facilitate its application in sustainable energy production.
  
  
  RESULTS
  The genomic analysis revealed that there are 154 genes encoding putative carbohydrate-active enzymes (CAZy) in P. ananatis Sd-1. This number is higher than that of compared cellulolytic and ligninolytic bacteria as well as other eight P. ananatis strains. The CAZy in P. ananatis Sd-1 contains a complete repertoire of enzymes required for cellulose and hemicellulose degradation. In addition, P. ananatis Sd-1 also possesses plenty of genes encoding potential ligninolytic relevant enzymes, such as multicopper oxidase, catalase/hydroperoxidase, glutathione S-transferase, and quinone oxidoreductase. Quantitative real-time PCR analysis of parts of genes encoding lignocellulolytic enzymes revealed that they were significantly up-regulated (at least P < 0.05) in presence of rice straw. Further identification of secretome of P. ananatis Sd-1 by nano liquid chromatography-tandem mass spectrometry confirmed that considerable amounts of proteins involved in lignocellulose degradation were only detected in rice straw cultures. Rice straw saccharification levels by the secretome of P. ananatis Sd-1 reached 129.11 ± 2.7 mg/gds. Correspondingly, the assay of several lignocellulolytic enzymes including endoglucanase, exoglucanase, β-glucosidase, xylanase-like, lignin peroxidase-like, and laccase-like activities showed that these enzymes were more active in rice straw relative to glucose substrates. The high enzymes activities were not attributed to bacterial cell densities but to the difference of secreted protein contents.
  
  
  CONCLUSIONS
  Our results indicate that P. ananatis Sd-1 can produce considerable lignocellulolytic enzymes including cellulases, hemicellulases, and ligninolytic relevant enzymes. The high activities of those enzymes could be efficiently induced by lignocellulosic biomass. This identified degradative system is valuable for the lignocellulosic bioenergy industry.
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Jiangshan
authors.initials: J
authors.lastname: Ma
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Keke
authors.initials: K
authors.lastname: Zhang
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Hongdong
authors.initials: H
authors.lastname: Liao
authors.affiliation: Department of Genetics, Institute for Plant Biotechnology, Stellenbosch University, Private Bag X1, Matieland, Stellenbosch, 7602 South Africa.|DNA Sequencing Unit, Central Analytical Facility, Stellenbosch University, Private Bag X1, Matieland, Stellenbosch, 7602 South Africa.
authors.forename: Stanton B
authors.initials: SB
authors.lastname: Hector
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Xiaowei
authors.initials: X
authors.lastname: Shi
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Jianglin
authors.initials: J
authors.lastname: Li
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Bin
authors.initials: B
authors.lastname: Liu
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Ting
authors.initials: T
authors.lastname: Xu
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Chunyi
authors.initials: C
authors.lastname: Tong
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Xuanming
authors.initials: X
authors.lastname: Liu
authors.affiliation: Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, College of Biology, Hunan University, Changsha, 410008 Hunan People's Republic of China.
authors.forename: Yonghua
authors.identifier: 0000-0002-0252-9464
authors.initials: Y
authors.lastname: Zhu
keywords: CAZy; Endophytic bacterium; Enzymes activities; Lignocellulose degradation; Pantoea ananatis Sd-1; Quantitative real-time PCR; Secretome
references.citation: Nat Prod Rep. 2011 Nov;28(12):1883-96
references.pmid: 21918777
references.citation: Syst Appl Microbiol. 2014 Feb;37(1):60-7
references.pmid: 24238986
references.citation: Proc Natl Acad Sci U S A. 2009 Feb 10;106(6):1954-9
references.pmid: 19193860
references.citation: Nat Protoc. 2006;1(6):2856-60
references.pmid: 17406544
references.citation: BMC Genomics. 2014 May 27;15:404
references.pmid: 24884520
references.citation: Sci Rep. 2015 Jun 12;5:10465
references.pmid: 26068013
references.citation: Nucleic Acids Res. 2012 Jul;40(Web Server issue):W445-51
references.pmid: 22645317
references.citation: J Bacteriol. 2012 Jan;194(2):534-5
references.pmid: 22207743
references.citation: Bioresour Technol. 2013 Jan;128:113-7
references.pmid: 23196230
references.citation: J Proteomics. 2012 Jun 27;75(12):3694-706
references.pmid: 22575269
references.citation: Methods. 2001 Dec;25(4):402-8
references.pmid: 11846609
references.citation: Biotechnol Biofuels. 2013 Oct 16;6(1):149
references.pmid: 24131596
references.citation: Mol Plant Pathol. 2009 May;10(3):325-35
references.pmid: 19400836
references.citation: Lett Appl Microbiol. 2014 Feb;58(2):123-9
references.pmid: 24111687
references.citation: Eukaryot Cell. 2012 Apr;11(4):482-93
references.pmid: 22345350
references.citation: Bioresour Technol. 2013 Dec;150:228-34
references.pmid: 24177155
references.citation: Appl Environ Microbiol. 2014 Aug;80(15):4692-701
references.pmid: 24837391
references.citation: Bioresour Technol. 2010 Oct;101(19):7592-7
references.pmid: 20537891
references.citation: Curr Opin Plant Biol. 2011 Aug;14(4):435-43
references.pmid: 21536480
references.citation: Appl Microbiol Biotechnol. 2015 Apr;99(7):2939-54
references.pmid: 25722022
references.citation: Nat Methods. 2011 Sep 29;8(10):785-6
references.pmid: 21959131
references.citation: Biochemistry. 2013 Feb 26;52(8):1437-45
references.pmid: 23356577
references.citation: ISME J. 2011 Aug;5(8):1323-31
references.pmid: 21368904
references.citation: Microbiol Res. 2010 Jul 20;165(5):392-401
references.pmid: 19853427
references.citation: Biochemistry. 2014 Jun 24;53(24):4047-58
references.pmid: 24870309
references.citation: FEBS Lett. 1993 May 24;323(1-2):135-40
references.pmid: 8495726
references.citation: Biochem J. 2004 Sep 15;382(Pt 3):769-81
references.pmid: 15214846
references.citation: Nucleic Acids Res. 2014 Jan;42(Database issue):D490-5
references.pmid: 24270786
references.citation: Curr Opin Chem Biol. 2014 Apr;19:1-7
references.pmid: 24780273
references.citation: J Proteome Res. 2010 Jun 4;9(6):3016-24
references.pmid: 20408570
references.citation: J Bacteriol. 2007 Sep;189(17):6260-5
references.pmid: 17601791
references.citation: Biotechnol Biofuels. 2013 Jan 08;6(1):1
references.pmid: 23298573
references.citation: J Ind Microbiol Biotechnol. 2009 Mar;36(3):319-32
references.pmid: 19137336
references.citation: Biosci Biotechnol Biochem. 2013;77(7):1474-80
references.pmid: 23832366
references.citation: J Ind Microbiol Biotechnol. 2008 May;35(5):377-91
references.pmid: 18338189
references.citation: Biotechnol Biofuels. 2013 Mar 21;6(1):41
references.pmid: 23514094
references.citation: J Appl Microbiol. 2007 Feb;102(2):410-5
references.pmid: 17241346
references.citation: PLoS One. 2009;4(4):e5271
references.pmid: 19384422
references.citation: Appl Environ Microbiol. 2005 Aug;71(8):4672-8
references.pmid: 16085862
references.citation: BMC Genomics. 2013 Apr 23;14:274
references.pmid: 23617724
references.citation: Curr Opin Biotechnol. 2011 Jun;22(3):394-400
references.pmid: 21071202
references.citation: Front Microbiol. 2013 Sep 19;4:280
references.pmid: 24065962
references.citation: FEMS Microbiol Rev. 2008 May;32(3):501-21
references.pmid: 18371173
references.citation: Biotechnol Lett. 2015 Nov;37(11):2279-88
references.pmid: 26209031
references.citation: Sci Rep. 2015 Feb 02;5:8161
references.pmid: 25641069
references.citation: J Ind Microbiol Biotechnol. 2014 Jan;41(1):1-7
references.pmid: 24212472
references.citation: Biosci Biotechnol Biochem. 1993 Oct;57(10):1655-9
references.pmid: 7764263
references.citation: Nucleic Acids Res. 2011 Apr;39(8):3240-54
references.pmid: 21227922
references.citation: Biotechnol Biofuels. 2012 Jul 02;5(1):45
references.pmid: 22747961
references.citation: Appl Environ Microbiol. 2011 Jun;77(12):4042-54
references.pmid: 21498747
references.citation: Microbiol Mol Biol Rev. 2002 Sep;66(3):506-77, table of contents
references.pmid: 12209002
references.citation: Gene. 2004 Aug 18;338(1):1-14
references.pmid: 15302401
references.citation: Appl Microbiol Biotechnol. 2012 Jan;93(2):723-41
references.pmid: 22159660