Molecular engineering of PETase for efficient PET biodegradation.

The widespread utilization of polyethylene terephthalate (PET) has caused a variety of environmental and health problems. Compared with traditional thermomechanical or chemical PET cycling, the biodegradation of PET may offer a more feasible solution. Though the PETase from Ideonalla sakaiensis (IsPETase) displays interesting PET degrading performance under mild conditions; the relatively low thermal stability of IsPETase limits its practical application. In this study, enzyme-catalysed PET degradation was investigated with the promising IsPETase mutant HotPETase (HP). On this basis, a carbohydrate-binding module from Bacillus anthracis (BaCBM) was fused to the C-terminus of HP to construct the PETase mutant (HLCB) for increased PET degradation. Furthermore, to effectively improve PET accessibility and PET-degrading activity, the truncated outer membrane hybrid protein (FadL) was used to expose PETase and BaCBM on the surface of E. coli (BL21with) to develop regenerable whole-cell biocatalysts (D-HLCB). Results showed that, among the tested small-molecular weight ester compounds (p-nitrophenyl phosphate (pNPP), p-Nitrophenyl acetate (pNPA), 4-Nitrophenyl butyrate (pNPB)), PETase displayed the highest hydrolysing activity against pNPP. HP displayed the highest catalytic activity (1.94 µM(p-NP)/min) at 50 °C and increased longevity at 40 °C. The fused BaCBM could clearly improve the catalytic performance of PETase by increasing the optimal reaction temperature and improving the thermostability. When HLCB was used for PET degradation, the yield of monomeric products (255.7 µM) was ∼25.5 % greater than that obtained after 50 h of HP-catalysed PET degradation. Moreover, the highest yield of monomeric products from the D-HLCB-mediated system reached 1.03 mM. The whole-cell catalyst D-HLCB displayed good reusability and stability and could maintain more than 54.6 % of its initial activity for nine cycles. Finally, molecular docking simulations were utilized to investigate the binding mechanism and the reaction mechanism of HLCB, which may provide theoretical evidence to further increase the PET-degrading activities of PETases through rational design. The proposed strategy and developed variants show potential for achieving complete biodegradation of PET under mild conditions.

[Immobilizing engineered Escherichia coli cells into zeolitic imidazolate framework 8 for efficient biosynthesis of Ala-Gln].

The α-amino acid ester acyltransferase (SAET) from Sphingobacterium siyangensis is one of the enzymes with the highest catalytic ability for the biosynthesis of l-alanyl-l-glutamine (Ala-Gln) with unprotected l-alanine methylester and l-glutamine. To improve the catalytic performance of SAET, a one-step method was used to rapidly prepare the immobilized cells (SAET@ZIF-8) in the aqueous system. The engineered Escherichia coli (E. coli) expressing SAET was encapsulated into the imidazole framework structure of metal organic zeolite (ZIF-8). Subsequently, the obtained SAET@ZIF-8 was characterized, and the catalytic activity, reusability and storage stability were also investigated. Results showed that the morphology of the prepared SAET@ZIF-8 nanoparticles was basically the same as that of the standard ZIF-8 materials reported in literature, and the introduction of cells did not significantly change the morphology of ZIF-8. After repeated use for 7 times, SAET@ZIF-8 could still retain 67% of the initial catalytic activity. Maintained at room temperature for 4 days, 50% of the original catalytic activity of SAET@ZIF-8 could be retained, indicating that SAET@ZIF-8 has good stability for reuse and storage. When used in the biosynthesis of Ala-Gln, the final concentration of Ala-Gln reached 62.83 mmol/L (13.65 g/L) after 30 min, the yield reached 0.455 g/(L·min), and the conversion rate relative to glutamine was 62.83%. All these results suggested that the preparation of SAET@ZIF-8 is an efficient strategy for the biosynthesis of Ala-Gln.

Current advances in the structural biology and molecular engineering of PETase.

Poly(ethylene terephthalate) (PET) is a highly useful synthetic polyester plastic that is widely used in daily life. However, the increase in postconsumer PET as plastic waste that is recalcitrant to biodegradation in landfills and the natural environment has raised worldwide concern. Currently, traditional PET recycling processes with thermomechanical or chemical methods also result in the deterioration of the mechanical properties of PET. Therefore, it is urgent to develop more efficient and green strategies to address this problem. Recently, a novel mesophilic PET-degrading enzyme (IsPETase) from Ideonella sakaiensis was found to streamline PET biodegradation at 30°C, albeit with a lower PET-degrading activity than chitinase or chitinase-like PET-degrading enzymes. Consequently, the molecular engineering of more efficient PETases is still required for further industrial applications. This review details current knowledge on IsPETase, MHETase, and IsPETase-like hydrolases, including the structures, ligand‒protein interactions, and rational protein engineering for improved PET-degrading performance. In particular, applications of the engineered catalysts are highlighted, including metabolic engineering of the cell factories, enzyme immobilization or cell surface display. The information is expected to provide novel insights for the biodegradation of complex polymers.

Fast jet proper motion discovered in a blazar at z=4.72.

High-resolution observations of high-redshift (z>4) radio quasars offer a unique insight into jet kinematics at early cosmological epochs, as well as constraints on cosmological model parameters. Due to the general weakness of extremely distant objects and the apparently slow structural changes caused by cosmological time dilation, only a couple of high-redshift quasars (HRQs) have been studied with parsec-scale resolutions, and with limited number of observing epochs. Here we report on very long baseline interferometry (VLBI) observations of a high-redshift blazar J1430 + 4204 (z=4.72) in the 8 GHz frequency band at five different epochs spanning 22 years. The source shows a compact core-jet structure with two jet components being identified within 3 milli-arcsecond (mas) scale. The long time span and multiple-epoch data allow for the kinematic studies of the jet components. That results in a jet proper motion of µ(J1) = 0.017 ±â€¯0.002 mas a-1 and µ(J2) = 0.156 ±â€¯0.015 mas a-1, respectively. For the fastest-moving outer jet component J2, the corresponding apparent transverse speed is (19.5±1.9)c. The inferred bulk jet Lorentz factor Γ=14.6±3.8 and viewing angle θ=2.2°±1.6° indicate highly relativistic beaming. The Lorentz factor and apparent proper motion are the highest measured to date among the z>4 jetted radio sources, while the jet kinematics is still consistent with the cosmological interpretation of quasar redshifts.

Evolving parsec-scale radio structure in the most distant blazar known.

Blazars are a sub-class of quasars with Doppler boosted jets oriented close to the line of sight, and thus efficient probes of supermassive black hole growth and their environment, especially at high redshifts. Here we report on Very Long Baseline Interferometry observations of a blazar J0906 + 6930 at z = 5.47, which enabled the detection of polarised emission and measurement of jet proper motion at parsec scales. The observations suggest a less powerful jet compared with the general blazar population, including lower proper motion and bulk Lorentz factor. This coupled with a previously inferred high accretion rate indicate a transition from an accretion radiative power to a jet mechanical power based transfer of energy and momentum to the surrounding gas. While alternative scenarios could not be fully ruled out, our results indicate a possibly nascent jet embedded in and interacting with a dense medium resulting in a jet bending.