Cloning and functional expression of α-galactosidase cDNA from Penicillium janczewskii zaleski

Abstract

In recent years, α-galactosidase has been attracting more and more attention because of its potential applications in many aspects. Using reverse transcription-polymerase chain reaction and rapid amplification of cDNA ends, a full-length cDNA sequence composed of 2,439 bp was cloned from Penicillium janczewskii zaleski and was subcloned into pPICZαA and transformed into Pichia pastoris strain X-33. In a 10-L fermentor, the recombinant yeast expressed α-galactosidase with a yield of 254 U/mL by methanol induction for 120 h. The recombinant enzyme showed the optimal activity at 40• C and pH 5.2. The Km values of the recombinant enzyme using p-nitrophenyl-α-D-galactopyranoside (pNPG), melibiose, raffinose and stachyose as substrates were 1, 16, 17.8 and 5.3 mM, respectively. Vmax values were 227.3, 116.7, 104.8, and 80.6 µM/min using pNPG, melibiose, raffinose and stachyose as substrates, respectively. The α-galactosidase exhibited no sensitivity to various metal ions and ethylenediaminetetraacetic acid, and hydrolyzed melibiose, raffinose and stachyose with different levels of galactose release. The biochemical characteristics of the α-galactosidase suggest that the enzyme may have a prospective application in feed industry as an additive.

Proteomic Insights on the Metabolism of Penicillium janczewskii during the Biotransformation of the Plant Terpenoid Labdanolic Acid

“…In our previous study, even at static conditions, germination of conidia was only observed in three out of the eight fungal strains tested (Frija et al, 2011 ). Several studies support the catabolic potential of P. janczewskii , e.g., for producing secondary metabolites (Madi and Katan, 1998 ), exo-inulinases (Pessoni et al, 2007 ), xylanases (Terrasan et al, 2010 , 2016 , 2017 ), and galactosidases (Zhang et al, 2011 ), as well as its ecological relevance in modulating plant–fungi interactions, either antagonizing (Madi and Katan, 1998 ) or stimulating (Kwasna, 2004 ) important plant pathogenic fungi.…”

Identification and Characterization of a Thermostable GH36 α-Galactosidase from Anoxybacillus vitaminiphilus WMF1 and Its Application in Synthesizing Isofloridoside by Reverse Hydrolysis

“…Contrary to our results, the α-galactosidase from Penicillium sp. F63 CGMCC 1669 [ 16 ] and Penicillium janczewskii zaleski [ 27 ] had optimum activity in the acidic pH range. The neutral or weak alkaline pH form of α-galactosidase is suitable for the hydrolysis of soy milk, since an acidic pH leads to the deposition of soy protein and gives milk its sour taste [ 26 ].…”

Heterologous Expression of a Thermostable α-Galactosidase from Parageobacillus thermoglucosidasius Isolated from the Lignocellulolytic Microbial Consortium TMC7

“…In the present study, the theoretical molecular weight of T26GAL was determined to be 83.47 kDa, which is consistent with that of the previously reported GH36 family α-galactosidases, such as the 82.9 kDa α-galactosidase from Gibberella sp. F75 [ 29 ] and the 81.8 kDa α-galactosidase isolated from P. janczewskii zaleski [ 28 ]. Homology-modeling using Swiss-Model showed that the 3D structure of T26GAL was similar to the α-galactosidases from L. acidophilus and Thermotoga maritima .…”