A Fungal Alpha-Galactosidase from Pseudobalsamia microspora Capable of Degrading Raffinose Family Oligosaccharides

Abstract

An alpha-galactosidase was purified from Pseudobalsamia microspora (PMG) to 1224.1-fold with a specific activity of 11,274.5 units/mg by ion-exchange chromatography and gel filtration. PMG is a monomeric protein with a molecular mass of 62 kDa as determined by SDS-PAGE and by gel filtration. Chemical modification using N-bromosuccinimide (NBS) resulted in a complete abrogation of the activity of PMG, suggesting that Trp is an amino acid essential to its activity. The activity was strongly inhibited by Hg(2+), Cd(2+), Cu(2+), and Fe(3+) ions. Three inner peptide sequences for PMG were obtained by liquid chromatography-tandem mass spectrometry (LC-MS-MS) analysis. When 4-nitrophenyl α-D-glucopyranoside (pNPGal) was used as substrate, the optimum pH and temperature of PMG were 5.0 and 55 °C, respectively. The Michaelis constant (K m) value of the alpha-galactosidase on pNPGal was 0.29 mM, and the maximal velocity (V max) was 0.97 μmol ml(-1) min(-1). Investigation by thin-layer chromatography (TLC) demonstrated its ability to hydrolyze raffinose and stachyose. Hence, it can be exploited in degradation of non-digestible oligosaccharides from food and feed industries.

A novel α-galactosidase from the thermophilic probiotic Bacillus coagulans with remarkable protease-resistance and high hydrolytic activity

“…Ag + and Hg 2+ completely inhibited enzyme activity, whereas the other metal ions had no evident effect on Aga-BC7050, including Cu 2+ and Fe 3+ . In contrast, many other α-galactosidases are partially or entirely inhibited by Cu 2+ and Fe 3+ [ 9 , 16 , 41 ]. No loss in α-galactosidase activity was observed when chemical reagents were present, except for SDS, which completely inhibited activity.…”

“…No obvious loss in activity was observed when the metal chelating agent EDTA was present, indicating that Aga-BC7050 is not a metal-based enzyme [ 16 ]. Aga-BC7050 was completely inactivated at an NBS concentration from 0.1 to 10 mM, which indicates a tryptophan residue is located in the active site and is indispensable for Aga-BC7050 enzymatic activity [ 9 ]. This is reminiscent of the finding that α-galactosidase from a strain of B .…”

“…α-Galactosidases also have utility in sugar production, where they can hydrolyze the raffinose from beet sugar syrups, increasing the output of crystallized sugar [ 7 ]. Fabry disease, an inherited lysosomal storage disease of glycosphingolipid metabolism caused by deficient or absent α-galactosidase A activity [ 8 ], is mitigated by the introduction of α-galactosidases [ 9 ]. α-Galactosidases can also be applied to convert blood from group B to group O by removing the terminal galactose residues from glycans on the surface of B-type red blood cells [ 10 ].…”

Galactomannan degradation by thermophilic enzymes: a hot topic for biotechnological applications

“…Thermophilic α-galactosidases from bacteria and fungi are attractive candidates due to their efficient catalytic activity and high stability under harsh conditions (Sarmiento et al 2015). Recently, several thermophilic enzymes have been discovered and characterized, such as the bacterial α-galactosidases from Neosarotrya fischeri P1 (Wang et al 2014), Bacillus megaterium VHM1 (Patil et al 2010), Bacillus coagulans (Zhao et al 2018) as well as the fungal α-galactosidases from Lenzites elegans (Sampietro et al 2012), Talaromyces leycettanus JCM12802 (Wang et al World Journal of Microbiology and Biotechnology _#####################_ 1 3 _####_ Page 4 of 12 2016), Pseudobalsamia microspore (Yang et al 2015a) and Rhizopus sp. F78 (Cao et al 2007) (Table 1).…”

Biochemical characteristics of a novel protease from the basidiomycete Amanita virgineoides

“…that Ca 2+ , Fe 2+ , and K + , in general, inhibited enzyme activity. However, the inhibition effects were less severe when concentrations of metal ions were further elevated . Mg 2+ , Ca 2+ , and K + ions, in this study, promoted protease activity and their most suitable concentrations were clearly shown in Table .…”