title: Direct amplification of new cellulase genes from woodland soil purified DNA.
abstract: Eight genes encoding cellulolytic enzymes were obtained by direct PCR amplification of genomic DNA recovered from woodland soil samples. The direct amplifications were carried out by using primers designed from available online cellulase nucleotide sequences. The isolated genes were all different from each other and homologous to endo-β-1,4-glucanases of Bacillus subtilis. The cellulases were functionally expressed in Escherichia coli and tested on soluble substrate at 37 and 60 °C, showing different cellulolytic activities. Among these, the enzyme renamed CelWS6 exhibited good activity at higher temperatures. Further analysis of CelWS6 showed a high performance in acid environments (between pH 4.0 and 6.0) and at elevated temperatures with its maximum activity at pH 5.0 and 50 °C. At the optimum pH, it was very stable since more than 80 % of its original activity was maintained after an incubation of 120 min at 60 °C. Because the cellulases had different cellulolytic activities, but similar amino acid sequences, it was possible to assess the relationship between sequence and protein function.
authors: Plantechno s.r.l, via Staffolo 60, 26041, Vicomoscano, Cremona, Italy.
authors: Marco
authors: M
authors: Cucurachi
authors: Matteo
authors: M
authors: Busconi
authors: Mariangela
authors: M
authors: Marudelli
authors: Giovanna
authors: G
authors: Soffritti
authors: Corrado
authors: C
authors: Fogher
mesh_terms: D044602:Cellulases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004247:DNA / Q000235:genetics*; D004789:Enzyme Activation; D004795:Enzyme Stability; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012987:Soil / Q000737:chemistry*; D013816:Thermodynamics
chemical_list: D012987:Soil; D004247:DNA; D044602:Cellulases
references: Mol Biol Rep. 2010 Apr;37(4):1923-9
references: 19669599
references: Appl Microbiol Biotechnol. 2011 Feb;89(4):1083-92
references: 20938774
references: Biochem J. 2012 Jan 1;441(1):95-104
references: 21880019
references: PLoS One. 2010 Jan 21;5(1):e8812
references: 20098679
references: Appl Environ Microbiol. 1982 Apr;43(4):777-80
references: 7081984
references: FEMS Microbiol Lett. 2008 May;282(1):44-51
references: 18355282
references: Biotechnol Biofuels. 2009 May 18;2:10
references: 19450243
references: Curr Opin Biotechnol. 2005 Oct;16(5):577-83
references: 16154338
references: Appl Microbiol Biotechnol. 2009 Mar;82(4):671-9
references: 19050861
references: Appl Microbiol Biotechnol. 2010 Feb;85(5):1241-9
references: 19908036
references: Science. 2011 Jan 28;331(6016):463-7
references: 21273488
references: Nat Rev Genet. 2008 Jun;9(6):433-43
references: 18487988
references: FEMS Microbiol Lett. 2013 Feb;339(2):93-101
references: 23181595
references: J Biotechnol. 2006 Oct 20;126(1):26-36
references: 16584799
references: Appl Environ Microbiol. 2011 Apr;77(7):2282-91
references: 21317267
references: Indian J Biochem Biophys. 2010 Aug;47(4):203-10
references: 21174947
references: Mol Biotechnol. 2008 Oct;40(2):195-201
references: 18576142
references: Eur J Biochem. 2001 Dec;268(24):6498-507
references: 11737205
references: Environ Microbiol. 2005 Dec;7(12):1996-2010
references: 16309396
references: J Phys Chem B. 2009 Apr 9;113(14):4970-7
references: 19292431