Li-Li Miao

The characteristics of a novel heterotrophic nitrification-aerobic denitrification bacterium, Bacillus methylotrophicus strain L7.

Bacillus methylotrophicus strain L7, exhibited efficient heterotrophic nitrification-aerobic denitrification ability, with maximum NH(4)(+)-N and NO(2)(-)-N removal rate of 51.58 mg/L/d and 5.81 mg/L/d, respectively. Strain L7 showed different gaseous emitting patterns from those strains ever described. When (15)NH(4)Cl, or Na(15)NO(2), or K(15)NO(3) was used, results of GC-MS indicated that N(2)O was emitted as the intermediate of heterotrophic nitrification or aerobic denitrification, while GC-IRMS results showed that N(2) was produced as end product when nitrite was used. Single factor experiments suggested that the optimal conditions for heterotrophic nitrification were sodium succinate as carbon source, C/N 6, pH 7-8, 0 g/L NaCl, 37 °C and a wide range of NH(4)(+)-N from 80 to 1000 mg/L. Orthogonal tests showed that the optimal conditions for aerobic denitrification were C/N 20, pH 7-8, 10 g/L NaCl and DO 4.82 mg/L (shaking speed 50 r/min) when nitrite was served as substrate.

Gene cloning and characterization of a cold-adapted β-glucosidase belonging to glycosyl hydrolase family 1 from a psychrotolerant bacterium Micrococcus antarcticus.

The gene bglU encoding a cold-adapted β-glucosidase (BglU) was cloned from Micrococcus antarcticus. Sequence analysis revealed that the bglU contained an open reading frame of 1419 bp and encoded a protein of 472 amino acid residues. Based on its putative catalytic domains, BglU was classified as a member of the glycosyl hydrolase family 1 (GH1). BglU possessed lower arginine content and Arg/(Arg+Lys) ratio than mesophilic GH1 β-glucosidases. Recombinant BglU was purified with Ni2+ affinity chromatography and subjected to enzymatic characterization. SDS-PAGE and native staining showed that it was a monomeric protein with an apparent molecular mass of 48 kDa. BglU was particularly thermolabile since its half-life time was only 30 min at 30°C and it exhibited maximal activity at 25°C and pH 6.5. Recombinant BglU could hydrolyze a wide range of aryl-β-glucosides and β-linked oligosaccharides with highest activity towards cellobiose and then p-nitrophenyl-β-d-glucopyranoside (pNPG). Under the optimal conditions with pNPG as substrate, the K(m) and k(cat) were 7 mmol/L and 7.85 × 103/s, respectively. This is the first report of cloning and characterization of a cold-adapted β-glucosidase belonging to GH1 from a psychrotolerant bacterium.

Marinobacter strain NNA5, a newly isolated and highly efficient aerobic denitrifier with zero N2O emission

An efficient aerobic denitrification bacterium, strain NNA5, was isolated and identified as Marinobacter sp. NNA5. NNA5 did not perform heterotrophic nitrification. GC/IRMS analysis revealed that (15)N2 was produced from Na(15)NO2 and K(15)NO3. GC/MS and quantitative analyses showed that no N2O emission occurred when nitrite or nitrate was used as substrate. Single factor experiments indicated that optimal conditions for aerobic denitrification were: sodium succinate or sodium pyruvate as carbon source, temperature 35 °C, NaCl concentration 2-4%, C/N ratio 6-8, pH 7.5, rotation speed 150 rpm (giving dissolved oxygen concentration 6.08 mg/L), NO3(-)-N concentration ranging from 140 to 700 mg/L. NNA5 displayed highly efficient aerobic denitrifying ability, with maximal NO3(-)-N removal rate 112.8 mg/L/d. In view of its ability to perform aerobic denitrification with zero N2O emission, NNA5 has great potential for future application in aerobic denitrification processes in industrial and aquaculture wastewater treatment systems.

Prokaryotic community structure driven by salinity and ionic concentrations in plateau lakes of the Tibetan plateau

ABSTRACT The prokaryotic community composition and diversity and the distribution patterns at various taxonomic levels across gradients of salinity and physiochemical properties in the surface waters of seven plateau lakes in the Qaidam Basin, Tibetan Plateau, were evaluated using Illumina MiSeq sequencing. These lakes included Lakes Keluke (salinity, <1 g/liter), Qing (salinity, 5.5 to 6.6 g/liter), Tuosu (salinity, 24 to 35 g/liter), Dasugan (salinity, 30 to 33 g/liter), Gahai (salinity, 92 to 96 g/liter), Xiaochaidan (salinity, 94 to 99 g/liter), and Gasikule (salinity, 317 to 344 g/liter). The communities were dominated by Bacteria in lakes with salinities of <100 g/liter and by Archaea in Lake Gasikule. The clades At12OctB3 and Salinibacter, previously reported only in hypersaline environments, were found in a hyposaline lake (salinity, 5.5 to 6.6 g/liter) at an abundance of ∼1.0%, indicating their ecological plasticity. Salinity and the concentrations of the chemical ions whose concentrations covary with salinity (Mg2+, K+, Cl−, Na+, SO4 2−, and Ca2+) were found to be the primary environmental factors that directly or indirectly determined the composition and diversity at the level of individual clades as well as entire prokaryotic communities. The distribution patterns of two phyla, five classes, five orders, five families, and three genera were well predicted by salinity. The variation of the prokaryotic community structure also significantly correlated with the dissolved oxygen concentration, pH, the total nitrogen concentration, and the PO4 3− concentration. Such correlations varied depending on the taxonomic level, demonstrating the importance of comprehensive correlation analyses at various taxonomic levels in evaluating the effects of environmental variable factors on prokaryotic community structures. Our findings clarify the distribution patterns of the prokaryotic community composition in plateau lakes at the levels of individual clades as well as whole communities along gradients of salinity and ionic concentrations.

Molecular Structural Basis for the Cold Adaptedness of the Psychrophilic β-Glucosidase BglU in Micrococcus antarcticus

ABSTRACT Psychrophilic enzymes play crucial roles in cold adaptation of microbes and provide useful models for studies of protein evolution, folding, and dynamic properties. We examined the crystal structure (2.2-Å resolution) of the psychrophilic β-glucosidase BglU, a member of the glycosyl hydrolase 1 (GH1) enzyme family found in the cold-adapted bacterium Micrococcus antarcticus. Structural comparison and sequence alignment between BglU and its mesophilic and thermophilic counterpart enzymes (BglB and GlyTn, respectively) revealed two notable features distinct to BglU: (i) a unique long-loop L3 (35 versus 7 amino acids in others) involved in substrate binding and (ii) a unique amino acid, His299 (Tyr in others), involved in the stabilization of an ordered water molecule chain. Shortening of loop L3 to 25 amino acids reduced low-temperature catalytic activity, substrate-binding ability, the optimal temperature, and the melting temperature (Tm ). Mutation of His299 to Tyr increased the optimal temperature, the Tm , and the catalytic activity. Conversely, mutation of Tyr301 to His in BglB caused a reduction in catalytic activity, thermostability, and the optimal temperature (45 to 35°C). Loop L3 shortening and H299Y substitution jointly restored enzyme activity to the level of BglU, but at moderate temperatures. Our findings indicate that loop L3 controls the level of catalytic activity at low temperatures, residue His299 is responsible for thermolability (particularly heat lability of the active center), and long-loop L3 and His299 are jointly responsible for the psychrophilic properties. The described structural basis for the cold adaptedness of BglU will be helpful for structure-based engineering of new cold-adapted enzymes and for the production of mutants useful in a variety of industrial processes at different temperatures.

Complete Genome Sequence of Rhodococcus sp. Strain IcdP1 Shows Diverse Catabolic Potential

ABSTRACT The complete genome sequence of Rhodococcus sp. strain IcdP1 is presented here. This organism was shown to degrade a broad range of high-molecular-weight polycyclic aromatic hydrocarbons and organochlorine pesticides. The sequence data can be used to predict genes for xenobiotic biodegradation and metabolism.