Alpha-amylase production by extremely halophilic archaeon Halococcus strain GUVSC8

Abstract

Starch hydrolyzing isolate GUVSC8 is obtained from commercial salt crystals of Vedaranyam, Tamil Nadu, India. GUVSC8 is categorized as an extreme halophile based on its growth on complex medium containing 25 percent (w/v) NaCl. Morphological, chemotaxonomic, and 16S rRNA gene sequencing reveal the cells to be coccus, producing bright orange C-50 bacterioruberin pigmentation with 95.95 percent similar to Halococcus hamelinensis DQ017835. The strain GUVSC8 grown in complex medium devoid of starch does not exhibit amylolytic activity, thereby confirming the induction of amylase production in presence of starch. On incubation of the starch with crude enzyme, numerous pores are observed on the starch granules after 24 h of incubation, indicating vigorous degradation and thereby confirming the amylolytic activity. Thin-layer chromatography (TLC) and liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) analysis reveal that the major end product obtained after amylolytic activity are glucose, maltose, maltotriose, maltotetrose, maltopentose, and other maltooligosaccharides, thus confirming it to be an alpha-amylase. Effects of NaCl, pH, and temperature, on activity of partially purified amylase, reveal best amylase activity at 2M NaCl, pH6, and 45 degrees C. The amylase is stable and active in presence of divalent cations such as Mg sup(2+), Ni sup(2+), Zn sup(2+), Ca sup(2+), and Co sup(2+). Enzyme activity significantly reduces with Cu sup(2+) and increases in presence of Mn sup(2+). The amylase is characterized as alpha amylase enzyme as it lost 87 percent of its activity in presence of EDTA. To the best of the author's knowledge, this is the first report on alpha-amylase production by salt crystal isolate belonging to the genus Halococcus.