Abstract.
An intracellular glucoamylase (E.C. 3.2.1.3) was purified to homogeneity from Lactobacillus amylovorus on a Fast Protein Liquid Chromatography System (FPLC) with a Mono Q ion-exchanger and two Superose 12 gel filtration columns arranged in series. The enzyme activity was quantified with a specific, chromogenic substrate, p-nitrophenyl-β-maltoside. Preparative gel electrophoresis was then used to further purify active enzyme fractions. Native polyacrylamide gel electrophoresis (Native-PAGE) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the purified enzyme showed a single protein band of molecular weight 47 kDa. Glucoamylase activity of the purified protein was confirmed by its ability to degrade starch on a 0.025% starch-polyacrylamide gel stained with I2/KI. Glucoamylase exhibited optimum catalytic activity at pH 6.0 and 45°C, and the enzyme had an isoelectric point near 4.39. The glucoamylase contained high levels of hydrophilic amino acids, comparable to fungal glucoamylases.
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Received: 12 July 1996 / Accepted: 10 September 1996
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James, J., Berger, JL. & Lee, B. Purification of Glucoamylase from Lactobacillus amylovorus ATCC 33621 . Curr Microbiol 34 , 186 –191 (1997). https://doi.org/10.1007/s002849900166
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DOI: https://doi.org/10.1007/s002849900166