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A novel endoglucanase gene (1425 bp), designated thrcel5A, was cloned from Thermoactinospora rubra YIM 77501Tand determined to be a member of glycoside hydrolase family 5. The putative amino acid sequence displayed 76% conservation with reported endoglucanases (GenBank: SCG57304.1) from Micromonospora siamensis. Thrcel5A was expressed in Escherichia coli BL 21 (DE3) and purified using Ni2+-affinity chromatography. The resulting purified protein displayed high hydrolytic activity against the sodium salt of carboxymethyl cellulose and β-(1, 3; 1, 4)-glucans from barley and beechwood xylan, with specific activities of 85.7 ± 1.5, 120.3 ± 2.6, and 22.9 ± 1.1 U/mg, respectively. The optimal pH and temperature for the recombinant enzyme were determined to be 8.5 and 60 °C, respectively. Additionally, ThrCel5A was thermotolerant as it retained more than 60% of its original activity after an incubation at 60 °C for 2 h. Moreover, ThrCel5A can hydrolyze β-(1, 3; 1, 4)-glucan into prebiotics, such as cellobiose, cellotriose, and cellotetrose. Its endoglucanase activity was significantly affected by link sequences and CBM2. Due to being an alkali-tolerant, thermostable, and multifunctional cellulolytic enzyme, ThrCel5A is an attractive candidate for use in production of prebiotics.

A β-glucosidase obtained from Thermobifida halotolerans YIM 90462T was expressed in Escherichia coli BL21 and subsequently characterized. The recombinant enzyme ThBGL1A showed optimal activity at 45 °C and pH 6, but also showed high activity from 40 to 55 °C and pH 5.6–6.6. Its half-life activity was 58 min at 50 °C. ThBGL1A exhibited notable cold-adapted activity, retaining 13.1%, 49.4%, and 83.5% of its optimal activity at 5, 25, and 30 °C, respectively. Kinetic characterization revealed an enzymatic turnover (Kcat) of 30 s−1 (cellobiose), 41.8 s−1 (p-nitrophenyl-β-d-glucopyranoside), and 52.6 s−1 (p-nitrophenyl-β-d-galactopyranoside). Moreover, ThBGL1A had high tolerance for salt, xylose, and glucose, which are extremely desirable features for industrial applications. Interestingly, its Ki for glucose was 932 mM and more than 80% of its optimal activity in the presence of 2000 mM xylose. After the addition of ThBGL1A (0.05 mg/ml) to a commercial cellulase reaction system, glucose yields from sugarcane bagasse were increased 20% and 18% after 1 day at 30 °C and 45 °C, respectively. Overall, this work identifies a cold-adapted, salt- and glucose-tolerant β-glucosidase with potential applications in commercial cellulose utilization and the bioenergy industry.

A gene encoding a β-xylosidase (designated as Thxyl43A) was cloned from strain Thermobifida halotolerans YIM 90462T. The open reading frame of this gene encodes 550 amino acid residues. The gene was over-expressed in Escherichia coli and the recombinant protein was purified. The monomeric Thxyl43A protein presented a molecular mass of 61.5 kDa. When p-nitrophenyl-β-d-xylopyranoside was used as the substrate, recombinant Thxyl43A exhibited optimal activity at 55 °C and pH 4.0 to 7.0, being thermostable by maintaining 47% of its activity after 30 h incubation at 55 °C. The recombinant enzyme retained more than 80% residual activity after incubation at pH range of 4.0 to 12.0 for 24 h, respectively, which indicated notable thermostability and pH stability of Thxyl43A. Moreover, Thxyl43A displayed high catalytic activity (> 60%) in presence of 5–35% NaCl (w/v) or 1–20% ionic liquid (w/v) or 1–50 mM xylose. These properties suggest that Thxyl43A has potential for promoting hemicellulose degradation and other industrial applications.

A xylanase gene (TrXyn10) from Thermoactinospora rubra YIM 77501T was cloned and expressed in Escherichia coli. The amino acid sequence displayed 78% homology with Microbispora mesophila xylanase (WP_062413927.1). The recombinant xylanase (TrXyn10), with MW 46.1 kDa, could hydrolyse beechwood, birchwood and oatspelt xylan. Based on the sequence, enzymatic properties and tertiary structure of the protein, TrXyn10 belongs to glycoside hydrolase family 10 (GH10). The optimal pH and temperature for the recombinant enzyme were determined to be 7.0 and 55 °C, respectively. TrXyn10 was stable over a wide pH range, and it retained more than 45% of the total activity at pH 6.0–12.0 for 12 h. In addition, the activity was greatly promoted, by approximately 200% of the initial activity, after incubation at pH 6.0 and 7.0 for 12 h. Based on enzymatic properties and product analysis, we showed that TrXyn10 is a neutral endoxylanase.

An alkalitolerant, thermotolerant and Gram-stain negative bacterium, designated strain YIM 78007T, was isolated from an alkaline geothermal soil sample from Hehua hot spring, Tengchong, Yunnan province, south-west China. Cells of strain YIM 78007T were observed to be aerobic and short rod-shaped. The colonies were observed to be orange-red, convex and circular. 16S rRNA gene sequence-based phylogenetic analysis showed that strain YIM 78007T clustered with members of the genus Roseomonas (with similarities from 97.2 to 92.2 %). Optimal growth of strain YIM 78007 occurs at 40–50 °C and pH 8.0–10.0. The predominant ubiquinone was identified as Q-10 and the major fatty acids were identified as C18:1 ω7c and C16:0. The polar lipids were identified as diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, two unidentified aminolipids and one unknown phospholipid. The G + C content of the genomic DNA was determined to be 63 mol %. The levels of DNA–DNA hybridization relatedness between strain YIM 78007T and its closet neighbours (Roseomonas lacus JCM 13283T and Roseomonas terrae JCM 14592T) were well below the threshold required for the proposal of a novel species. The results of physiological and biochemical characteristics, the phylogenetic analysis, as well as low DNA–DNA hybridization values, allowed the phenotypic and genotypic differentiation of strain YIM 78007T from its closest phylogenetic neighbours. Therefore, strain YIM 78007T is considered to represent a novel species of the genus Roseomonas, for which the name Roseomonas alkaliterrae sp. nov. is proposed. The type strain is YIM 78007T (=BCRC 80644T = JCM 19656T).

An extracellular xylanase from halophilic Streptomonospora sp. YIM 90494 was purified to homogeneity from a fermentation broth by ammonium sulphate precipitation, gel filtration chromatography and ion exchange chromatography. The purified xylanase appeared as a single protein band on SDS-PAGE with a molecular mass of approximately 50 kDa. The xylanase had maximum activity at pH 7.5 and 55 °C. The enzyme was stable over a broad pH range (pH 4.0–10.0) and showed good thermal stability when being incubated at 60 °C for 2 h. Kinetic experiments indicated that the enzyme had K m and V max values of 19.24 mg/mL and 6.1 μmol/min/mg, respectively, using birch wood xylan as substrate. The inhibitory effects of various metal ions and chemical agents on the xylanase activity were investigated. It is greatly interesting to note that Ag+ ion and SDS, which strongly inhibited most xylanases reported previously increases the xylanase activity in this study. These characteristics suggest that the enzyme with new properties has considerable potential in industrial applications.

A Gram-stain negative aerobic bacterium, designated YIM 77924(T), was isolated from a geothermally heated soil sample collected at Rehai National Park, Tengchong, Yunnan province, south-west China. Growth was found to occur from 55 to 75 °C (optimum 65 °C), pH 6.0-8.0 (optimum pH 7.0) and 0-1 % NaCl (w/v). Cells were observed to be rod-shaped and the colonies convex, circular, smooth, yellow and non-transparent. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain YIM 77924(T) belongs to the genus Thermus. The 16S rRNA gene sequence similarity values between strain YIM 77924(T) and other species of the genus Thermus were all below 97 %. The polar lipids of strain YIM 77924(T) were determined to be aminophospholipid, phospholipid and glycolipid. The predominant respiratory quinone was determined to be MK-8 and the G+C content was 66.64 mol%. The major fatty acids identified were iso-C(16:0), iso-C(15:0), iso-C(17:0) and C(16:0). On the basis of the morphological and chemotaxonomic characteristics as well as genotypic data, strain YIM 77924(T) is proposed to represent a novel species, Thermus tengchongensis sp. nov., in the genus Thermus. The type strain is YIM 77924(T) (=KCTC 32025(T) = CCTCC AB2012063(T)).

A Gram-negative and aerobic bacterium, designated YIM 77875(T), was isolated from a geothermal soil sample collected at Rehai National Park, Tengchong, Yunnan Province, south-west China. Bacterial growth occurred from 37 to 65 °C (optimum 50 °C), pH 6.0-8.0 (optimum pH 7.0) and 0-1 % NaCl (w/v). Cells were rod-shaped and colonies were convex, circular, smooth, yellow and non-transparent. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain YIM 77875(T) belongs to the genus Lysobacter. The 16S rRNA gene sequence similarity values between strain YIM 77875(T) and other species of the genus Lysobacter were all below 94.7 %. The polar lipids of strain YIM 77875(T) were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and five unknown phospholipids. The predominant respiratory quinone was Q-8 and the G+C content was 68.8 mol%. Major fatty acids were iso-C(16:0), iso-C(15:0) and iso-C(11:0). On the basis of the morphological and chemotaxonomic characteristics, as well as genotypic data, strain YIM 77875(T) represents a novel species, Lysobacter thermophilus sp. nov., in the genus Lysobacter. The type strain is YIM 77875(T) (CCTCC AB 2012064(T) = KCTC 32020(T)).