Glucoamylase isoform (GAII) purified from a thermophilic fungus Scytalidium thermophilum 15.8 with biotechnological potential

Abstract

Scytalidium thermophilum 15.8 produced two extracellular glucoamylases. Using a DEAE-Cellulose chromatographic column glucoamylases form II (GAII) was separated and purified from glucoamylases form I (GAI) that was previously purified and characterised (Cereia et al., 2000) when the filtrate of the culture medium was applied to a DEAE-Cellulose chromatographic column. GAII bound to the DEAECellulose and was eluted with a NaCl gradient, while GAI did not bind to the resin. GAII presented
electrophoretic homogeneity in 6% denaturing and non-denaturing PAGE, separately, with a molecular mass of 83 kDa, after the second round DEAE-Cellulose purification step. The enzyme pI was 7.2.
Optima pH and activity temperature were 5.5 and 55ºC respectively for starch and maltose as substrates, with a termostability of 2.5 min at 60ºC. Enzymatic activities were activated by 1 mM Na+, Mn2+ and Mg2+ or 10 mM NH4+, Ba2+ and Mg2+. The carbohydrate content was 10%. The kinetic parameters Km and Vmax with starch and maltose as substrate were 0.2 and 1.5 mg/ml, and 22.3 and 4.39 U/mg of protein, respectively. The amino acid sequence of GAII had 92% homology with the
glucoamylase of Humicola grisea var. thermoidea after 13 cycles. Generally, GAII had different properties compared with GAI (Cereia et al., 2000).