Extra-cellular glucoamylase of Humicola sp. produced under submerged growth condition (10.44 U mg(-1) protein) was purified to homogeneity level by using three-step purification procedure. Crude enzyme was subjected to ammonium sulphate precipitation, Hiload anion exchange chromatography and hydrophobic interaction column chromatography on FPLC purification system. The purified glucoamylase was monomeric in nature because native molecular mass on gel column chromatography and sub-unit mass on SDS-PAGE were the same, i.e. 72.8 kDa. Activation energy for soluble starch hydrolysis was 21.09 kJ mol(-1), while temperature quotient (Q(10)) was 1.01. The enzyme was stable over a pH range of 3.5-5.9 and gave pH optimum of 4.7. Temperature optimum of the glucoamylase was 55 degrees C. V-max for soluble starch hydrolysis was 56.63 U mg(-1) protein and K-m was 0.26 mg (soluble starch) ml(-1). The turn over (k(cat)) was 69 s(-1). The pKa(1) and pKa(2) of ionizable groups of active site controlling V-max were 3.65 and 6.3, respectively. Thermodynamic parameters for soluble starch hydrolysis were as follows: Delta H* = 18.36 kJ mol(-1), Delta G* = 69.06 kJ mol(-1), Delta S* = -154.56 J mol(-1) K-1, Delta G(E-S)* = -3.71 kJ mol(-1) and Delta G(E-T)* = -26.41 kJ mol(-1). Thermodynamic parameters (Delta H*, Delta G*, Delta S*) for irreversible inactivation of glucoamylase at different temperatures (45-55 degrees C) were also determined. Current report has novelty as it explained for the first time kinetics and thermodynamics of soluble starch hydrolysis and irreversible inactivation of glucoamylase from Humicola sp. (C) 2007 Elsevier Inc. All rights reserved.