Culture filtrates of Cladosporium resinae ATCC 20495 contain a mixture of enzymes able to convert starch and pullulan efficiently into D-glucose. Culture conditions for optimal production of the pullulan-degrading activity have been established. The amylolytic enzyme preparation was fractionated by ion-exchange and molecular-sieve chromatography, and shown to contain alpha-D-glucosidase, alpha-amylase, and two glucoamylases. The glucoamylases have been purified to homogeneity and their substrate specificities investigated. One of the glucoamylases (termed P) readily hydrolyses the (1 leads to 6)-alpha-D linkages in pullulan, amylopectin, isomaltose, panose, and 6(3)-alpha-D-glucosylmaltotriose. Each of the glucoamylases cleaves the (1 leads to 6)-alpha-D linkage in panose much more readily than that in isomaltose.