Substrate specificity of the α- l-arabinofuranosidase from Rhizomucor pusillus HHT-1

Abstract

The α- l-arabinofuranosidase (AF) from the fungus Rhizomucor pusillus HHT-1 released arabinose at appreciable rates from (1→5)-α- l-arabinofuranooligosaccharides, sugar beet arabinan and debranched arabinan. This enzyme preferentially hydrolyzed the terminal arabinofuranosyl residue [α-(1→5)-linked] of the arabinan backbone rather than the arabinosyl side chain [α-(1→3)-linked residues]. The enzyme-hydrolyzed arabinan reacted at and debranched the arabinan almost at the same rate, and the degree of conversion for both cases was 65%. Methylation analysis of arabinan showed that the arabinosyl-linkage proportions were 2:2:2:1, respectively, for (1→5)-Ara f, T-Ara f, (1→3, 5)-Ara f and (1→3)-Ara f, while the ratios for the AF-digested arabinan shifted to 3:1:2:1. Enzyme digestion resulted in an increase in the proportion of (1→5)-linked arabinose and a decrease in the proportion of terminal arabinose indicated this AF cleaved the terminal arabinosyl residue of the arabinan back bone [α-(1→5)-linked residues]. Peak assignments in the 13C NMR spectra also confirmed this linkage composition of four kinds of arabinose residues. Both 1H and 13C NMR spectra are dominated by signals of the α-anomeric configuration of the arabinofuranosyl moieties. No signals were recorded for arabinopyranosyl moieties in the NMR spectra. Methylation and NMR analysis of native and AF-digested arabinan revealed that this α- l-arabinofuranosidase can only hydrolyse α- l-arabinofuranosyl residues of arabinan.