Enzymatic Characterization of a Rumen Microorganism-Derived Multifunctional Glycoside Hydrolase and Its GH26 Domain with Mannanase Activity

In this study, a novel multifunctional glycoside hydrolase (GH) with two distinct domains homologous to the GH family 5 (GH5) and family 26 (GH26) was isolated from the rumen microorganism <i>Segatella bryantii</i>. The heterologous expression product of this enzyme exhibited both endo-β-1,4-glucanase and endo-β-1,4-mannanase activities. Intriguingly, segmental expression studies indicated that the GH26 domain alone contributed to the β-mannanase activity, and its specific activity reached 2060 U/mg under optimal conditions (30 °C, pH 5.5). Furthermore, site-directed mutagenesis confirmed that the glutamic acid residues at positions 165 and 276 were indispensable for the catalytic activity of the GH26 domain. Collectively, a novel multifunctional GH from a symbiotic microorganism of ruminants was identified. Preliminary enzymatic characterizations of its GH26 family domain, which has independent β-mannanase activities, were determined.