A Novel Glycosyl Hydrolase Family 16 ��-Agarase from the Agar- Utilizing Marine Bacterium Gilvimarinus agarilyticus JEA5: the First Molecular and Biochemical Characterization of Agarase in Genus Gilvimarinus

Abstract

The agarase gene was identified from a draft genome sequence of JEA5, an agar-utilizing marine bacterium. Recently, three agarase-producing bacteria, ,, and , in the genus were reported. However, there have been no reports of the molecular characteristics and biochemical properties of these agarases. In this study, we analyzed the molecular characteristics and biochemical properties of agarases in . Gaa16A comprised a 1,323-bp open reading frame encoding 441 amino acids. The predicted molecular mass and isoelectric point were 49 kDa and 4.9, respectively. The amino acid sequence of Gaa16A showed features typical of glycosyl hydrolase family 16 (GH16) β-agarases, including a GH16 domain, carbohydrate-binding region (RICIN domain), and signal peptide. Recombinant Gaa16A (excluding the signal peptide and carbohydrate-binding region, rGaa16A) was expressed as a fused protein with maltose-binding protein at its N-terminus in . rGaa16A had maximum activity at 55°C and pH 7.0 and 103 U/mg of specific activity in the presence of 2.5 mM CaCl₂. The enzyme hydrolyzed agarose to yield neoagarotetraose as the main product. This enzyme may be useful for industrial production of functional neoagaro-oligosaccharides.

A Novel Agarase, Gaa16B, Isolated from the Marine Bacterium Gilvimarinus agarilyticus JEA5, and the Moisturizing Effect of Its Partial Hydrolysis Products

“…We previously reported a draft genome sequence for the agar-degrading bacterium G. agarilyticus JEA5 [21], and we were the first to describe the molecular characteristics and biochemical properties of a β-agarase (Gaa16A) isolated from G. agarilyticus JEA5 [14]. Here, we describe Gaa16B, a novel neoagarotriose-producing β-agarase.…”

“…β-agarases are classified into four families based on their amino acid sequence homology: glycoside hydrolase 16 (GH16), GH50, GH86, and GH118 [4]. To date, many β-agarases have been found in various genera, including Agarivorans [5], Alteromonas [6], Bacillus [7], Catenovulum [8], Pseudoalteromonas [9], Pseudomonas [10], Vibrio [11], Streptomyces [12], Saccharophagus [13], and Gilvimarinus [14].…”

A Novel Neoagarotriose-Producing Agarase from the Marine Bacterium Gilvimarinus Agarilyticus JEA5

“…We have previously reported a draft genome sequence for the agar degrading bacterium G. agarilyticus JEA5 [18], and were the rst to describe the molecular characteristics and biochemical properties of a βagarase (Gaa16A) isolated from G. agarilyticus JEA5 [19]. Here, we describe Gaa16B, a novel neoagarotriose-producing β-agarase.…”

“…The Gaa16B amino acid sequence showed the highest identity and similarity with a carbohydrate binding protein from G. polysacchariticus (WP_049721028.1), which has not yet been characterized. The NCBI database contains three genomes from the Gilvimarinus genus; however, only one agarase (Gaa16A from G. agarilyticus JEA5) has been characterized thus far [19]. Compared to the amino acid sequences of characterized agarases, Gaa16B displayed the highest similarity to an agarase from Saccharophagus degradans 2-40 (with a sequence identity and similarity of only 55.9% and 70.9%, respectively).…”

“…The β-agarases are classi ed into four families on the basis of their amino acid sequence homology: glycoside hydrolase 16 (GH16), GH50, GH86, and GH118 [4]. To date, many β-agarases have been found from many different genera including Agarivorans [5], Alteromonas [6], Bacillus [7], Catenovulum [8], Pseudoalteromonas [9], Pseudomonas [10], Vibrio [11], Streptomcyes [12], Saccharophagus [13] and Gilvimarinus [14].…”