Characterization of Sphingobacterium sp. Ab3 Lipase and Its Coexpression with LEA Peptides.

Sphingobacterium sp. is a yellowish Gram-negative bacterium that is usually characterized by high concentrations of sphingophospholipids as lipid components. As microbial enzymes have been in high demand in industrial fields in the past few decades, this study hopes to provide significant information on lipase activities of Sphingobacterium sp., since limited studies have been conducted on the Sphingobacterium sp. lipase. A microbe from one collected Artic soil sample, ARC4, was identified as psychrotolerant Sphingobacterium sp., and it could grow in temperatures ranging from 0°C to 24°C. The expression of Sphingobacterium sp. lipase was successfully performed through an efficient approach of utilizing mutated group 3 late embryogenesis abundant (G3LEA) proteins developed from Polypedilum vanderplanki. Purified enzyme was characterized using a few parameters, such as temperature, pH, metal ion cofactors, organic solvents, and detergents. The expressed enzyme is reported to be cold adapted and has the capability to work efficiently under neutral pH (pH 5.0 to 7.0), cofactors like Na+ ion, and the water-like solvent methanol. Addition of nonionic detergents greatly enhanced the activity of purified enzyme. IMPORTANCE The mechanism of action of LEA proteins has remained unknown to many; in this study we reveal their presence and improved protein expression due to the molecular shielding effect reported by others. This paper should be regarded as a useful example of using such proteins to influence an existing expression system to produce difficult-to-express proteins.

Bioprospecting and Molecular Identification of Used Transformer Oil-Degrading Bacteria for Bioplastics Production.

One of the major impediments to the commercialization of biodegradable plastic is the high cost of substrate. Consequently, there is a continuous search for effective microorganisms and cheaper carbon substrates to reduce the high production cost. In this study, waste transformer oil-degrading bacteria were isolated from soil, wastewater, and sediment samples, using a mineral salt medium (MSM) supplemented with 1% waste transformer oil as the sole carbon source. The isolates were screened for polyhydroxyalkanoates (PHA) production using Nile red staining and fluorescence microscopy. PHA granules accumulation was confirmed using transmission electron microscopy. Oil degradation analysis was accomplished using solvent extraction and gravimetric methods whereas, the bacteria were identified using 16S DNA sequence homology. A total of 62 transformer oil-degrading bacteria were isolated, out of which 16 (26%) showed positive results for Nile red fluorescence microscopy. The identified organisms belong to four different taxonomic genera of Acinetobacter, Bacillus, Proteus, and Serratia. The percentage of oil degradation observed among the different isolates ranged between 19.58% and 57.51%. Analysis of the PHA extracted from the selected isolate revealed the presence of medium chain length polyhydroxyalkanoates (mcl-PHA). The findings of this work have further highlighted the diversity of the bacteria capable of utilizing waste streams such as waste transformer oil. Consequently, the isolates can be explored as agents of converting waste transformer oil into bioplastics.

Purification and Characterization of a Thermostable Lipase from Geobacillus thermodenitrificans IBRL-nra.

Thermostable lipase from Geobacillus thermodenitrificans IBRL-nra was purified and characterized. The production of thermostable lipase from Geobacillus thermodenitrificans IBRL-nra was carried out in a shake-flask system at 65°C in cultivation medium containing; glucose 1.0% (w/v); yeast extract 1.25% (w/v); NaCl 0.45% (w/v) olive oil 0.1% (v/v) with agitation of 200 rpm for 24 hours. The extracted extracellular crude thermostable lipase was purified to homogeneity by using ultrafiltration, Heparin-affinity chromatography, and Sephadex G-100 gel-filtration chromatography by 34 times with a final yield of 9%. The molecular weight of the purified enzyme was estimated to be 30 kDa after SDS-PAGE analysis. The optimal temperature for thermostable lipase was 65°C and it retained its initial activity for 3 hours. Thermostable lipase activity was highest at pH 7.0 and stable for 16 hours at this pH at 65°C. Thermostable lipase showed elevated activity when pretreated with BaCl(2), CaCl(2), and KCl with 112%, 108%, and 106%, respectively. Lipase hydrolyzed tripalmitin (C16) and olive oil with optimal activity (100%) compared to other substrates.