Agarase Production by Marine Pseudoalteromonas sp. MHS: Optimization, and Purification

Abstract

Agar is an essential polysaccharide that has been utilized in numerous fields. Many kinds of literature have been published regarding agarolytic microorganisms’ isolation and agarases biochemical studies. In this search, a local marine agarolytic bacterium associated with marine alga Ulva lactuca surface was isolated and identified as Pseudoalteromonas sp. MHS. The agarase production was parallel to the growth of Pseudoalteromonas sp. MHS as cells displayed a lag phase (2 h), subsequently an exponential growth that prolonged till 10 h where maximum growth (OD550nm = 3.9) was achieved. The enzyme activity increased rapidly as cells increased exponentially where the maximum activity of 0.22 U/mL was achieved after 8h and remained constant till 12 h during the stationary phase of growth. Agarase production was optimized using Plackett-Burman statistical design by measuring enzyme activity as a response and the design was validated using a verification experiment; the activity of the enzyme increased from 0.22 U/mL to 0.29 U/mL. Pseudoalteromonas sp. MHS agarase was partially purified and its molecular weight (MW) was determined by SDSPAGE (15-25 kDa). Agarase showed approximately 94% of its activity at 40 °C. The enzyme stability decreased as the temperature increased; the enzyme could retain about 98, 90, 80, 75, and 60% of its activity at 20, 30, 40, 50, and 60 °C, respectively. Biomass of the red alga Pterocladia capillacea proved to be a suitable substrate for agarase production using Pseudoalteromonas sp. MHS; the enzyme activity recorded after 24 h of incubation was 0.35 U/mL compared to 0.29 U/mL from the optimized medium.