Apple-pectin hairy regions were prepared from apple pectin by combined action of the recombinant Aspergillus niger enzymes endopolygalacturonase II and pectin methylesterase and the A. tubigensis exopolygalacturonase. Using this enzymically prepared pectin fraction, an additional activity of the A. tubigensis exopolygalacturonase was discovered only when the substrate was chemically saponified and when D-galacturonate, a potent inhibitor of the enzyme, was removed from the incubation mixture. The new reaction product was purified and could be hydrolysed by A. niger beta-xylosidase into D-galacturonate and beta-D-xylose in a 1:1 ratio, which identified it as xylogalacturonate. The results demonstrate that exopolygalacturonase is not only active on galacturonan but also on xylogalacturonan. The enzyme thus accomodates a substrate in which the terminal galacturonic acid residue carries a single xylose substitution. The well-defined substrate specificity of exopolygalacturonase opens the possibility for use of this enzyme in biotechnological applications, such as preparing pectins that are methylated at the non-reducing end, and for studying the fine structure of xylogalacturonan in pectin.