Expression and characterization of a pectin methylesterase from Aspergillus niger ZJ5 and its application in fruit processing
Section snippets
Reagents, strains and media
Substrate citrus pectin, citrus pectin with 77% esterified (DE77) and citrus pectin with 85% esterified (DE85) were purchased from Sigma–Aldrich (San Diego, CA, USA). The pEASY-T3 vector, Escherichia coli Trans1-T1 competent cells and Fastpfu DNA polymerase (TransGen, Beijing, China) were used for gene cloning and sequencing. The plasmid pPIC9 and P. pastoris GS115 (Invitrogen, Carlsbad, CA, USA) were used to construct the expression vector and for heterologous expression. T4-DNA ligase and
Cloning and sequence analysis of pme-zj5a
Full-length DNA and cDNA corresponding to the pme-zj5a gene (GenBank accession number: MG397069) (Supplementary Fig. S1) were cloned from A. niger ZJ5 by PCR using specific primers. The DNA was 1088 bp in length and included two introns. Its cDNA was 984 bp long and encoded a polypeptide of 327 amino acids and a stop codon. The deduced PME-ZJ5A protein contained a catalytic domain and a putative N-terminal signal peptide (residues 1–19) of carbohydrate esterase family 8 of Carbohydrate-Active
Discussion
More than 200 amino acid sequences of PMEs from fungal were collected in the National Center for Biotechnology Information database (http://www.ncbi.nlm.nih.gov/protein). The majority of them have a mesophilic temperature optimum (40–60 °C) and an acidic pH optimum, such as PME from A. flavus whose optimal activity occurs at 55 °C and pH 4.8 (25), PME from A. repens maximum activity occurred at 30 °C and pH 6.5 (26), and recombinant PME from A. niger showed maximum activity at pH 4.7 and 50 °C
Acknowledgments
This research was supported by the National High Technology Research and Development Program of China (2012AA022207) and the National Natural Science Foundation of China (grant number 31402007 and 31201313). We thank Angie Deng (Molecular Physiology and Biophysics, Vanderbilt University) for her valuable comments during manuscript writing and revising.
The authors declare no competing financial interest.
References (39)
- et al.
A novel low-temperature-active pectin methylesterase from Penicillium chrysogenum F46 with high efficiency in fruit firming
Food Chem.
(2014) - et al.
Microbial pectinolytic enzymes: a review
Process Biochem.
(2005) - et al.
Food-processing enzymes from recombinant microorganisms - a review
Regul. Toxicol. Pharmacol.
(2006) - et al.
High hydrostatic pressure protection of a pectinase cocktail against thermal inactivation
J. Food Eng.
(2013) - et al.
New insights into pectin methylesterase structure and function
Trends Plant Sci.
(2007) - et al.
High-yield production of a low-temperature-active polygalacturonase for papaya juice clarification
Food Chem.
(2013) - et al.
Influence of calcium on pectin methylesterase behaviour in the presence of medium methylated pectins
Carbohydr. Polym.
(2011) - et al.
Applications of pectinases in the commercial sector: a review
Bioresour. Technol.
(2001) - et al.
Characterization and expression of a genomic pectin methyl esterase-encoding gene in Aspergillus niger
Gene
(1991) - et al.
Efficient bioconversion of lactose in milk and whey: immobilization and biochemical characterization of a beta-galactosidase from the dairy Streptococcus thermophilus LMD9 strain
Res. Microbiol.
(2010)
Separation and characterization of three beta-galactosidases from Bacillus circulans
Biochim. Biophys. Acta
Taxonomic structure of the yeasts and lactic acid bacteria microbiota of pineapple (Ananas comosus L. Merr.) and use of autochthonous starters for minimally processing
Food Microbiol.
Fungal biotechnology in food and feed processing
Food Res. Int.
Potential application of pectinase in developing functional foods
Annu. Rev. Food Sci. Technol.
Enzymatic added extraction and clarification of fruit juices-A review
Crit. Rev. Food Sci.
Apple juice clarification by immobilized pectolytic enzymes in packed or fluidized bed reactors
J. Agric. Food Chem.
Constitutive expression, purification and characterisation of pectin methylesterase from Aspergillus niger in Pichia pastoris for potential application in the fruit juice industry
J. Sci. Food Agric.
Enzymatic modification of a model homogalacturonan with the thermally tolerant pectin methylesterase from citrus: 1. nanostructural characterization, enzyme mode of action, and effect of pH
J. Agric. Food Chem.
Partially esterified oligogalacturonides are the preferred substrates for pectin methylesterase of Aspergillus niger
Biochem. J.
Cited by (0)
- ‡
The first two authors contributed equally to this work.