Biochemical Characterization of a New β-Agarase from Cellulophaga algicola

Abstract

Cellulophaga algicola DSM 14237, isolated from the Eastern Antarctic coastal zone, was found to be able to hydrolyze several types of polysaccharide materials. In this study, a predicted β-agarase (CaAga1) from C. algicola was heterologously expressed in Escherichia coli. The purified recombinant CaAga1 showed specific activities of 29.39, 20.20, 14.12, and 8.99 U/mg toward agarose, pure agar, and crude agars from Gracilaria lemaneiformis and Porphyra haitanensis, respectively. CaAga1 exhibited an optimal temperature and pH of 40 oC and 7, respectively. CaAga1 was stable over a wide pH range from 4 to 11. The recombinant enzyme showed an unusual thermostability, that is, it was stable at temperature below or equal to 40oC and around 70 oC, but was thermolabile at about 50 oC. With the agarose as the substrate, the Km and Vmax values for CaAga1 were 1.19 mg/mL and 36.21 U/mg, respectively. The reducing reagent (dithiothreitol) enhanced the activity of CaAga1 by more than one fold. In addition, CaAga1 was salt-tolerant given that it retained approximately 70% of the maximum activity in the presence of 2 M NaCl. The thin layer chromatography results indicated that CaAga1 is an endo-type β-agarase and efficiently hydrolyzed agarose into neoagarotetraose (NA4) and neoagarohexaose (NA6). A structural model of CaAga1 in complex with neoagarooctaose (NA8) was built by homology modeling and explained the hydrolysis pattern of CaAga1.

Metagenomics analysis of cultured mucosal bacteria from colorectal cancer and adjacent normal mucosal tissues

“…baltica phages [36, 37]; however, some species in this genus can hydrolyse various polysaccharides (e.g. agar, starch, gelatin and carboxymethylcellulose) [38–40]. The findings of Myroides odoratimimus and Cellulophaga baltica in CRC tissues were obviously different with previous studies.…”

“…No relevant clinical reports have been published on this bacterium except for C. baltica phages [36,37]; however, some species in this genus can hydrolyse various polysaccharides (e.g. agar, starch, gelatin and carboxymethylcellulose) [38][39][40].…”

Enhanced Stability of β-Agarase Immobilized on Streptavidin-Coated Fe3O4 Nanoparticles: Effect of Biotin Linker Length

“…6 According to amino acid sequence similarity comparisons, β-agarases are mainly categorized into four glycoside hydrolase (GH) families: GH16, GH50, GH86, and GH118. 7 Interestingly, different GH families of β-agarases exhibit different protein sequences and agarose degradation patterns. β-Agarases of GH16, GH86, and GH118 family are endolytic enzyme and generate a series of even-numbered oligosaccharides; however, a few β-agarases of the GH50 family have been identified with an exomode of action, and the final products of GH50 β-agarases are relatively single, usually neoagarotetraose or neoagarobiose.…”

“…During the last decade, β-agarase have been isolated from marine sediments, marine algae, marine mollusks, sea water, fresh water, and soil . According to amino acid sequence similarity comparisons, β-agarases are mainly categorized into four glycoside hydrolase (GH) families: GH16, GH50, GH86, and GH118 . Interestingly, different GH families of β-agarases exhibit different protein sequences and agarose degradation patterns.…”

Molecular Characterization of a Novel 1,3-α-3,6-Anhydro-L-Galactosidase, Ahg943, with Cold- and High-Salt-Tolerance from Gayadomonas joobiniege G7

“…In addition, agar can be used as a supporting material for enzyme or bacterial immobilization to enhance the stability of the system, which allows long-term operation [ 6 , 7 ]. Agar can also be used to manufacture biodegradable polymers, such as bioplastics [ 8 ], and can be used in wet-fiber [ 9 ], eco-friendly biocleaning processes [ 10 ] as well as in medical treatments, such as microencapsulation [ 11 ], drug delivery [ 12 ], and bone generation [ 13 ]. In this sense, agar is expected to be widely used in the food and chemical industry as well as in the medical field.…”