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Molecular cloning and characterization of a novel α-galactosidase gene from Penicillium sp. F63 CGMCC 1669 and expression in Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
18188 42713 2007 8 PDF Available
Title
Molecular cloning and characterization of a novel α-galactosidase gene from Penicillium sp. F63 CGMCC 1669 and expression in Pichia pastoris
Abstract

An extracellular α-galactosidase (EC 3.2.1.22) from Penicillium sp. F63 CGMCC 1669, designated Agl1, was purified to homogeneity by means of fast protein liquid chromatography (FPLC) with a molecular mass of 82 kDa on SDS-PAGE while 330 kDa on native gradient PAGE. Based on the partial amino acid sequences of the purified protein, Agl1 gene encoding the α-galactosidase was cloned and sequenced. The ORF of Agl1 consisted of 2205 nucleotides encoding a protein of 735 amino acids including 21 residues of a putative signal peptide in its N-terminal. Agl1 was intron-less, and a GUG codon was considered the start codon of the gene. Agl1 possessed the highest sequence identity of 69.5% with α-galactosidase AglC from Aspergillus niger and showed a little homology to those of Penicillium purpurogenum and Penicillium simplicissimum reported previously, namely, 7.5% and 8.0%, respectively. The mature protein had been expressed extracellularly in Pichia pastoris with a yield of 111 U/ml in a 3-l fermenter. The optimum pH and temperature of the recombinant α-galactosidase were 5.0 and 40 °C, respectively. The recombinant α-galactosidase hydrolyzed the release of galactose from natural oligosaccharides, such as melibiose, raffinose, and stachyose. To our knowledge, this is the first report of gene cloning for Penicillium α-galactosidase belonging to family 36 of glycosyl hydrolases and expression in Pichia pastoris.

Keywords
α-Galactosidase; Penicillium sp.; Molecular cloning; Gene expression; Pichia pastoris
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Molecular cloning and characterization of a novel α-galactosidase gene from Penicillium sp. F63 CGMCC 1669 and expression in Pichia pastoris
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 40, Issue 5, 3 April 2007, Pages 1373–1380
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us