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Purification and characterization of a novel laccase from Cerrena sp. HYB07 with dye decolorizing ability.

Yang J, Lin Q, Ng TB, Ye X, Lin J - PLoS ONE (2014)

Bottom Line: Its gene and cDNA sequences were obtained.Putative cis-acting transcriptional response elements were identified in the promoter region.The high production yield and activity, robustness and dye decolorizing capacity make LacA and Cerrena sp.

View Article: PubMed Central - PubMed

Affiliation: College of Biological Sciences and Technology, Fuzhou University, Fuzhou, Fujian, China; National Engineering Laboratory for Enzyme Expression, Fuzhou, Fujian, China.

ABSTRACT
Laccases (EC 1.10.3.2) are a class of multi-copper oxidases with important industrial values. A basidiomycete strain Cerrena sp. HYB07 with high laccase yield was identified. After cultivation in the shaking flask for 4 days, a maximal activity of 210.8 U mL(-1) was attained. A 58.6-kDa laccase (LacA) with 7.2% carbohydrate and a specific activity of 1952.4 U mg(-1) was purified. 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) was the optimal substrate, with Km and kcat being 93.4 µM and 2468.0 s(-1), respectively. LacA was stable at 60°C, pH 5.0 and above, and in organic solvents. Metal ions Na+, K+, Ca2+, Mg2+, Mn2+, Zn2+ enhanced LacA activity, while Fe2+ and Li+ inhibited LacA activity. LacA decolorized structurally different dyes and a real textile effluent. Its gene and cDNA sequences were obtained. Putative cis-acting transcriptional response elements were identified in the promoter region. The high production yield and activity, robustness and dye decolorizing capacity make LacA and Cerrena sp. HYB07 potentially useful for industrial and environmental applications such as textile finishing and wastewater treatment.

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Promoter analysis of LacA gene.First nucleotide of the start codon ATG is designated as +1. Putative cis-acting responsive elements are underlined/boxed and labeled according to following abbreviations. ACE1: ACE1 copper-responsive element; ARE: antioxidant response element; CreA: CreA-binding site; HSE: heat shock response element; MRE: metal response element; NIT2: consensus sequences for binding of NIT2 transcription factor; PRE: putative response element; XRE: xenobiotic response element. Putative CCAAT and TATA boxes are underlined.
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pone-0110834-g006: Promoter analysis of LacA gene.First nucleotide of the start codon ATG is designated as +1. Putative cis-acting responsive elements are underlined/boxed and labeled according to following abbreviations. ACE1: ACE1 copper-responsive element; ARE: antioxidant response element; CreA: CreA-binding site; HSE: heat shock response element; MRE: metal response element; NIT2: consensus sequences for binding of NIT2 transcription factor; PRE: putative response element; XRE: xenobiotic response element. Putative CCAAT and TATA boxes are underlined.

Mentions: SiteFinding PCR was adopted to further extend the 5′-flanking region of LacA, rendering a region of 1,544 bp upstream of the start codon, referred to as the LacA promoter. Bioinformatics analysis revealed multiple putative cis-acting transcription regulation sites within the LacA promoter sequence in both orientations (Fig. 6). A TATA box was located 92 bp upstream from the start codon ATG, and three CCAAT boxes were found at positions −324, −405 and −1134. The LacA promoter contained two metal response elements (MREs) with the consensus sequence TGCRCNC at positions −1013 and −1227 and one xenobiotic response element (XRE) with the core sequence TNGCGTG [38] at −1223. Apart from MREs and XREs, LacA had many ACE1 copper-responsive transcription factor binding sites, consisting of the HWHNNGCTGD or NTNNHGCTGN core [39], at positions −11, −456, −774 and −1336, respectively. In addition, one antioxidant response element (ARE) adhering to the consensus sequence TGACNNNGC [40] was present at −1035 in the LacA promoter. Putative response elements (PREs) [38], often found in basidiomycete laccase promoter sequences, were also present within the 5′-flanking sequence of LacA: a TGGGT was located at position −1360, an inverted one at −1377, and two ATATC at −122 and −795. Furthermore, two heat shock response elements (HSEs) composed of alternately oriented NGAAN repeats [41] were found at −422 and −857 in the LacA promoter. LacA also contained multiple transcription factor binding sites involved in nitrogen and carbon regulation. Two putative CreA-binding sites (SYGGRG) were found at −648 and −706, and five NIT2 binding sites adhering to sequence TATCDH [5] were scattered at −120, −525, −793, −1139 and −1286. No stress response elements (STREs) with the consensus sequence of CCCCT or Sp-1 transcription factor recognition site (GGGCGG) [39] were identified in the LacA promoter sequence.


Purification and characterization of a novel laccase from Cerrena sp. HYB07 with dye decolorizing ability.

Yang J, Lin Q, Ng TB, Ye X, Lin J - PLoS ONE (2014)

Promoter analysis of LacA gene.First nucleotide of the start codon ATG is designated as +1. Putative cis-acting responsive elements are underlined/boxed and labeled according to following abbreviations. ACE1: ACE1 copper-responsive element; ARE: antioxidant response element; CreA: CreA-binding site; HSE: heat shock response element; MRE: metal response element; NIT2: consensus sequences for binding of NIT2 transcription factor; PRE: putative response element; XRE: xenobiotic response element. Putative CCAAT and TATA boxes are underlined.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4214704&req=5

pone-0110834-g006: Promoter analysis of LacA gene.First nucleotide of the start codon ATG is designated as +1. Putative cis-acting responsive elements are underlined/boxed and labeled according to following abbreviations. ACE1: ACE1 copper-responsive element; ARE: antioxidant response element; CreA: CreA-binding site; HSE: heat shock response element; MRE: metal response element; NIT2: consensus sequences for binding of NIT2 transcription factor; PRE: putative response element; XRE: xenobiotic response element. Putative CCAAT and TATA boxes are underlined.
Mentions: SiteFinding PCR was adopted to further extend the 5′-flanking region of LacA, rendering a region of 1,544 bp upstream of the start codon, referred to as the LacA promoter. Bioinformatics analysis revealed multiple putative cis-acting transcription regulation sites within the LacA promoter sequence in both orientations (Fig. 6). A TATA box was located 92 bp upstream from the start codon ATG, and three CCAAT boxes were found at positions −324, −405 and −1134. The LacA promoter contained two metal response elements (MREs) with the consensus sequence TGCRCNC at positions −1013 and −1227 and one xenobiotic response element (XRE) with the core sequence TNGCGTG [38] at −1223. Apart from MREs and XREs, LacA had many ACE1 copper-responsive transcription factor binding sites, consisting of the HWHNNGCTGD or NTNNHGCTGN core [39], at positions −11, −456, −774 and −1336, respectively. In addition, one antioxidant response element (ARE) adhering to the consensus sequence TGACNNNGC [40] was present at −1035 in the LacA promoter. Putative response elements (PREs) [38], often found in basidiomycete laccase promoter sequences, were also present within the 5′-flanking sequence of LacA: a TGGGT was located at position −1360, an inverted one at −1377, and two ATATC at −122 and −795. Furthermore, two heat shock response elements (HSEs) composed of alternately oriented NGAAN repeats [41] were found at −422 and −857 in the LacA promoter. LacA also contained multiple transcription factor binding sites involved in nitrogen and carbon regulation. Two putative CreA-binding sites (SYGGRG) were found at −648 and −706, and five NIT2 binding sites adhering to sequence TATCDH [5] were scattered at −120, −525, −793, −1139 and −1286. No stress response elements (STREs) with the consensus sequence of CCCCT or Sp-1 transcription factor recognition site (GGGCGG) [39] were identified in the LacA promoter sequence.

Bottom Line: Its gene and cDNA sequences were obtained.Putative cis-acting transcriptional response elements were identified in the promoter region.The high production yield and activity, robustness and dye decolorizing capacity make LacA and Cerrena sp.

View Article: PubMed Central - PubMed

Affiliation: College of Biological Sciences and Technology, Fuzhou University, Fuzhou, Fujian, China; National Engineering Laboratory for Enzyme Expression, Fuzhou, Fujian, China.

ABSTRACT
Laccases (EC 1.10.3.2) are a class of multi-copper oxidases with important industrial values. A basidiomycete strain Cerrena sp. HYB07 with high laccase yield was identified. After cultivation in the shaking flask for 4 days, a maximal activity of 210.8 U mL(-1) was attained. A 58.6-kDa laccase (LacA) with 7.2% carbohydrate and a specific activity of 1952.4 U mg(-1) was purified. 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) was the optimal substrate, with Km and kcat being 93.4 µM and 2468.0 s(-1), respectively. LacA was stable at 60°C, pH 5.0 and above, and in organic solvents. Metal ions Na+, K+, Ca2+, Mg2+, Mn2+, Zn2+ enhanced LacA activity, while Fe2+ and Li+ inhibited LacA activity. LacA decolorized structurally different dyes and a real textile effluent. Its gene and cDNA sequences were obtained. Putative cis-acting transcriptional response elements were identified in the promoter region. The high production yield and activity, robustness and dye decolorizing capacity make LacA and Cerrena sp. HYB07 potentially useful for industrial and environmental applications such as textile finishing and wastewater treatment.

Show MeSH
Related in: MedlinePlus