Effects of phosphonium-based ionic liquids on the lipase activity evaluated by experimental results and molecular docking.

In this work, the effect of several phosphonium-based ionic liquids (ILs) on the activity of lipase from Burkholderia cepacia (BCL) was evaluated by experimental assays and molecular docking. ILs comprising different cations ([P4444 ]+ , [P444(14) ]+ , [P666(14) ]+ ) and anions (Cl- , Br- , [Deca]- , [Phosp]- , [NTf2 ]- ) were investigated to appraise the individual roles of IL ions on the BCL activity. From the activity assays, it was found that an increase in the cation alkyl chain length leads to a decrease on the BCL enzymatic activity. ILs with the anions [Phosp]- and [NTf2 ]- increase the BCL activity, while the remaining [P666(14) ]-based ILs with the Cl- , Br- , and [Deca]- anions display a negative effect on the BCL activity. The highest activity of BCL was identified with the IL [P666(14) ][NTf2 ] (increase in the enzymatic activity of BCL by 61% at 0.055 mol·L-1 ). According to the interactions determined by molecular docking, IL cations preferentially interact with the Leu17 residue (amino acid present in the BCL oxyanion hole). The anion [Deca]- has a higher binding affinity compared to Cl- and Br- , and mainly interacts by hydrogen-bonding with Ser87, an amino acid residue which constitutes the catalytic triad of BCL. The anions [Phosp]- and [NTf2 ]- have high binding energies (-6.2 and -5.6 kcal·mol-1 , respectively) with BCL, and preferentially interact with the side chain amino acids of the enzyme and not with residues of the active site. Furthermore, FTIR analysis of the protein secondary structure show that ILs that lead to a decrease on the α-helix content result in a higher BCL activity, which may be derived from an easier access of the substrate to the BCL active site.

Petroleum hydrocarbon degradation by isolated mangrove bacteria / Degradación de hidrocarburos de petróleo por bacterias aisladas de manglares

The petroleum hydrocarbon contamination represents a worldwide problem, since its accumulation promotes a serious environmental impact. Thereby, the use of microorganisms, such as those from mangrove micro biota, as degrading agents of various carbon sources is poorly exploited in environmental remediation processes. Thus, this in vitro study evaluated the degrading potential of isolated bacteria from mangrove sediments in the degradation of petroleum hydrocarbons. Analysis of the genetic diversity using the 16S rRNA marker revealed closely related (99%) sequences with Proteobacterium, Pseudomonas and Exiguobacterium. Results showed the bacterial growth in the mineral saline medium (MSM) containing 1% petroleum or diesel, as carbon sources. This growth was determinated by optical density at 595 nm for 15 days, with sample withdrawal every 48 h. Bacterial growth indicated the hydrocarbon metabolization. However, bacteria were more efficient at degrading petroleum. Overall, experimental data displayed the potential application of these bacteria in bioremediation processes, due to their metabolic and adaptive capacities to grow in a rich hydrocarbon medium.

Los hidrocarburos de petróleo representan un problema mundial, pues su acumulación promueve un serio impacto ambiental. Así, el uso de microorganismos, por ejemplo los de la microbiota de manglares, como agentes degradadores de diversas fuentes de carbono, es poco explotado en procesos de remediación ambiental. Así, este estudio evaluó in vitro el potencial degradador de bacterias aisladas de sedimento de manglar en la degradación de hidrocarburos. El análisis genético usando el marcador 16S rRNA reveló secuencias íntimamente relacionadas (99%) con Proteobacterium, Pseudomonas y Exiguobacterium. Los resultados mostraron el crecimiento de bacterias en medio salino mineral (MSM) conteniendo petróleo o diesel al 1%, como fuentes de carbono. Este crecimiento, determinado por densidad óptica (DO) a 595 nm durante 15 días, con toma de muestras a cada 48 h, indicó la matabolización de hidrocarburos. Sin embargo, las bacterias fueron más eficientes en degradarlos. Por lo tanto, los resultados muestran la potencial aplicación de las bacterias en procesos de biorremediación por su capacidad metabólica y adaptativa de crecimiento usando hidrocarburos.

Immobilization and characterisation of a lipase from a new source, Bacillus sp. ITP-001.

A new source of lipase from Bacillus sp. ITP-001 was immobilized by physical adsorption on the polymer poly(3-hydroxybutyrate-co-hydroxyvalerate) (PHBV) in aqueous solution. The support and immobilized lipase were characterised, compared to the lyophilised lipase, with regard to the specific surface area, adsorption-desorption isotherms, pore volume (V(p)) and size (dp) by nitrogen adsorption, differential scanning calorimetry, thermogravimetric analysis, chemical composition analysis, Fourier transform infrared spectroscopy and biochemical properties. The immobilized enzyme displayed a shift in optimum pH towards the acidic side with an optimum at pH 4.0, whereas the optimum pH for the free enzyme was at pH 7.0; the optimum temperature of activity was 80 and 37 °C for the free and immobilized enzyme, respectively. The inactivation rate constant for the immobilized enzyme at 37 °C was 0.0038 h⁻¹ and the half-life was 182.41 h. The kinetic parameters obtained for the immobilized enzyme gave a Michaelis-Menten constant (K(m)) of 49.10 mM and a maximum reaction velocity (V(max)) of 205.03 U/g. Furthermore, the reuse of the lipase immobilized by adsorption allowed us to observe that it could be reused for 10 successive cycles, duration of each cycle (1 h), maintaining 33 % of the initial activity.

Immobilization of Candida rugosa lipase on poly(3-hydroxybutyrate-co-hydroxyvalerate): a new eco-friendly support.

The overall objective of this study is to evaluate the morphological [scanning electron microscopy (SEM)], physicochemical [differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), chemical composition analysis, Fourier-transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR)], and biochemical properties of Candida rugosa lipase (CRL) immobilized on a natural biopolymer poly(3-hydroxybutyrate-co-hydroxyvalerate) (PHBV) in aqueous solution. CRL was immobilized by physical adsorption with efficiency of 30%. Compared with free CRL enzyme, there were slight changes in immobilized CRL activity as a function of temperature (from 37°C to 45°C), but a similar optimal pH value of 7.0. Inactivation rate constants for immobilized CRL enzyme were 0.009 and 0.334 h⁻¹, and half-lives were 77 and 2 h at 40°C and 60°C, respectively. Kinetic parameters obtained for immobilized CRL include the Michaelis-Menten constant of K(m) = 213.18 mM and maximum reaction velocity of V(max) = 318.62 U/g. The operational stability of immobilized CRL was tested repeatedly, and after 12 cycles of reuse, the enzyme retained 50% activity. Based on our results, we propose that PHBV-immobilized CRL could serve as a promising biocatalyst in several industrial applications.