Isolation and purification of carrageenase enzyme from Pseudoalteromonas carrageenova IFO 12985 (Protobacterium)
Carrageenan is a gel-forming polysaccharide extracted from the cell wall of red seaweeds. One of the most common forms of this sulfated galactan is the t-carrageenan that is biodegraded by an anzyme called t-carrageenase. This extracellular enzyme specifically cleaves the alpha-1, 4 linkage between the D-galactopyranosyl-4-sulfate and 3,6- anhydro-alpha -D-galactopyranosyl-2-sulfate residues. Several studies have been conducted on the isolation, purification and characterization of t-carrageenase from various marine bacteria such as Zobella galactanovorans and Alteromonas fortis. Here, the authors report on the isolation, purification and partial characterization of t-carrageenase present in Pseudoalteromonas carrageenovora. From the cell-free supernatant of cultures grown on t-carrageenan, t-carrageenase was purified by ammonium sulfate precipitation and followed by ion-exchange chromatography on CM and DEAE. The isolated enzyme had a molecular weight of about 55 kDa as estimated by SDS-PAGE, in agreement with the reported MW for t-carrageenase. Hydrolytic activity was observed towards t-carrageenan and not against t-carrageenan when analyzed by the reducing sugar assay. Analysis of the unfractionated hydrolysate by sup13C NMR also confirmed that degradation had taken place.