40 research outputs found
Fungal bioremediation of polyethylene:challenges and perspectives
Plastics have become ubiquitous in both their adoption as materials and as environmental contaminants. Widespread pollution of these versatile, man-made, and largely petroleum-derived polymers has resulted from their long-term mass production, inappropriate disposal, and inadequate end of life management. Polyethylene (PE) is at the forefront of this problem, accounting for one third of plastic demand in Europe in part due to its extensive use in packaging (European Parliament, 2020). Current recycling and incineration processes do not represent sustainable solutions to tackle plastic waste, especially once it becomes littered, and the development of new waste-management and remediation technologies are needed. Mycoremediation (fungal-based biodegradation) of PE has been the topic of several studies over the last two decades. The utility of these studies is limited by an inconclusive definition of biodegradation and a lack of knowledge regarding the biological systems responsible. This review highlights relevant features of fungi as potential bioremediation agents, before discussing the evidence for fungal biodegradation of both high- and low-density PE. An up-to-date perspective on mycoremediation as a future solution to PE waste is provided. [Abstract copyright: This article is protected by copyright. All rights reserved.
The Lysozyme Inhibitor Thionine Acetate Is Also an Inhibitor of the Soluble Lytic Transglycosylase Slt35 from Escherichia coli
Lytic transglycosylases such as Slt35 from E. coli are enzymes involved in bacterial cell wall remodelling and recycling, which represent potential targets for novel antibacterial agents. Here, we investigated a series of known glycosidase inhibitors for their ability to inhibit Slt35. While glycosidase inhibitors such as 1-deoxynojirimycin, castanospermine, thiamet G and miglitol had no effect, the phenothiazinium dye thionine acetate was found to be a weak inhibitor. IC50 values and binding constants for thionine acetate were similar for Slt35 and the hen egg white lysozyme. Molecular docking simulations suggest that thionine binds to the active site of both Slt35 and lysozyme, although it does not make direct interactions with the side-chain of the catalytic Asp and Glu residues as might be expected based on other inhibitors. Thionine acetate also increased the potency of the beta-lactam antibiotic ampicillin against a laboratory strain of E. coli
Thermal Adaptation of Dihydrofolate Reductase from the Moderate ThermophileGeobacillus stearothermophilus
The thermal melting temperature of dihydrofolate reductase from Geobacillus stearothermophilus (BsDHFR) is 30 °C higher than that of its homologue from the psychrophile Moritella profunda. Additional proline residues in the loop regions of BsDHFR have been proposed to enhance the thermostability of BsDHFR, but site-directed mutagenesis studies reveal that these proline residues contribute only minimally. Instead, the high thermal stability of BsDHFR is partly due to removal of water-accessible thermolabile residues such as glutamine and methionine, which are prone to hydrolysis or oxidation at high temperatures. The extra thermostability of BsDHFR can be obtained by ligand binding, or in the presence of salts or cosolvents such as glycerol and sucrose. The sum of all these incremental factors allows BsDHFR to function efficiently in the natural habitat of G. stearothermophilus, which is characterized by temperatures that can reach 75 °C
Loop Interactions during Catalysis by Dihydrofolate Reductase fromMoritella profunda
Dihydrofolate reductase (DHFR) is often used as a model system to
study the relation between protein dynamics and catalysis. We have studied a
number of variants of the cold-adapted DHFR from Moritella profunda
(MpDHFR), in which the catalytically important M20 and FG loops have been
altered, and present a comparison with the corresponding variants of the wellstudied
DHFR from Escherichia coli (EcDHFR). Mutations in the M20 loop do not
affect the actual chemical step of transfer of hydride from reduced nicotinamide
adenine dinucleotide phosphate to the substrate 7,8-dihydrofolate in the catalytic
cycle in either enzyme; they affect the steady state turnover rate in EcDHFR but
not in MpDHFR. Mutations in the FG loop also have different effects on catalysis
by the two DHFRs. Despite the two enzymes most likely sharing a common catalytic cycle at pH 7, motions of these loops,
known to be important for progression through the catalytic cycle in EcDHFR, appear not to play a significant role in MpDHFR
Reclassification of the Specialized Metabolite Producer Pseudomonas mesoacidophila ATCC 31433 as a Member of the Burkholderia cepacia Complex.
Pseudomonas mesoacidophila ATCC 31433 is a Gram-negative bacterium, first isolated from Japanese soil samples, that produces the monobactam isosulfazecin and the β-lactam-potentiating bulgecins. To characterize the biosynthetic potential of P. mesoacidophila ATCC 31433, its complete genome was determined using single-molecule real-time DNA sequence analysis. The 7.8-Mb genome comprised four replicons, three chromosomal (each encoding rRNA) and one plasmid. Phylogenetic analysis demonstrated that P. mesoacidophila ATCC 31433 was misclassified at the time of its deposition and is a member of the Burkholderia cepacia complex, most closely related to Burkholderia ubonensis The sequenced genome shows considerable additional biosynthetic potential; known gene clusters for malleilactone, ornibactin, isosulfazecin, alkylhydroxyquinoline, and pyrrolnitrin biosynthesis and several uncharacterized biosynthetic gene clusters for polyketides, nonribosomal peptides, and other metabolites were identified. Furthermore, P. mesoacidophila ATCC 31433 harbors many genes associated with environmental resilience and antibiotic resistance and was resistant to a range of antibiotics and metal ions. In summary, this bioactive strain should be designated B. cepacia complex strain ATCC 31433, pending further detailed taxonomic characterization.IMPORTANCE This work reports the complete genome sequence of Pseudomonas mesoacidophila ATCC 31433, a known producer of bioactive compounds. Large numbers of both known and novel biosynthetic gene clusters were identified, indicating that P. mesoacidophila ATCC 31433 is an untapped resource for discovery of novel bioactive compounds. Phylogenetic analysis demonstrated that P. mesoacidophila ATCC 31433 is in fact a member of the Burkholderia cepacia complex, most closely related to the species Burkholderia ubonensis Further investigation of the classification and biosynthetic potential of P. mesoacidophila ATCC 31433 is warranted
Protein motions and dynamic effects in enzyme catalysis
The role of protein motions in promoting the chemical step of enzyme catalysed reactions remains a subject of considerable debate. Here, a unified view of the role of protein dynamics in dihydrofolate reductase catalysis is described. Recently the role of such motions has been investigated by characterising the biophysical properties of isotopically substituted enzymes through a combination of experimental and computational analyses. Together with previous work, these results suggest that dynamic coupling to the chemical coordinate is detrimental to catalysis and may have been selected against during DHFR evolution. The full catalytic power of Nature's catalysts appears to depend on finely tuning protein motions in each step of the catalytic cycle
Synthetic routes, characterization and photophysical properties of luminescent, surface functionalized nanodiamonds
The functionalization of small diameter (ca. 50 nm) polycarboxylated nanodiamond particles using amide coupling methodologies in both water and acetonitrile solvent has been investigated. In this manner, the surfaces of nanodiamond particles were adorned with different luminescent moieties, including a green fluorescent 1,8-naphthalimide species (Nap-1), and a red emitting ruthenium(II) tris-bipyridine complex (Ru-1), as well as dual functionalization with both luminophores. Comprehensive characterization of the surface functionalized nanodiamonds has been achieved using a combination of dynamic light scattering, nanoparticle tracking analysis, transmission electron microscopy, X-ray photoelectron spectroscopy, zeta potential measurements, microwave plasma atomic emission spectroscopy and time-resolved photophysics. The tendency of the functionalized nanodiamonds to aggregate reflects the degree of surface substitution, yielding small aggregates with typical particle sizes ca. 150 nm. This is likely to be driven by the reduction of the zeta potential, concomitant with the conversion of surface charged carboxylate groups to neutral amide functions. The results show that luminescent nanodiamond materials can be synthesised with tuneable photophysical properties
Chemoenzymatic Assembly of Isotopically Labeled Folates
Pterin-containing natural products have diverse functions in life but an efficient and easy scheme for their in vitro synthesis is not available. Here, we report a chemo-enzymatic 14-step, one-pot synthesis that can be used to generate 13C- and 15N-labeled dihydrofolates (H2F) from glucose, guanine and p-aminobenzoyl-L-glutamic acid. This synthesis stands out from previous approaches to produce H2F in that the average yield of each step is >91% and it requires only one single purification step. The use of a one-pot reaction allowed us to overcome potential problems with individual steps during the synthesis. The availability of labeled dihydrofolates allowed the measurement of heavy atom isotope effects for the reactions catalyzed by the drug target dihydrofolate reductase and established that protonation at the N5 position of H2F and hydride transfer to the C4 position occur in a stepwise mechanism. This chemo-enzymatic pterin synthesis can be applied to the efficient production of other folates and a range of other natural compounds with applications in nutritional, medical and cell biological research
TCR‐induced alteration of primary MHC peptide anchor residue
The HLA‐A*02:01‐restricted decapeptide EAAGIGILTV, derived from melanoma antigen recognized by T‐cells‐1 (MART‐1) protein, represents one of the best‐studied tumor associated T‐cell epitopes, but clinical results targeting this peptide have been disappointing. This limitation may reflect the dominance of the nonapeptide, AAGIGILTV, at the melanoma cell surface. The decapeptide and nonapeptide are presented in distinct conformations by HLA‐A*02:01 and TCRs from clinically relevant T‐cell clones recognize the nonapeptide poorly. Here, we studied the MEL5 TCR that potently recognizes the nonapeptide. The structure of the MEL5‐HLA‐A*02:01‐AAGIGILTV complex revealed an induced fit mechanism of antigen recognition involving altered peptide–MHC anchoring. This “flexing” at the TCR–peptide–MHC interface to accommodate the peptide antigen explains previously observed incongruences in this well‐studied system and has important implications for future therapeutic approaches. Finally, this study expands upon the mechanisms by which molecular plasticity can influence antigen recognition by T cells
昇温熱分解¹⁴C法による堆積物年代推定と北極海イベント層の層位学的研究 [論文内容及び審査の要旨]
The role of protein
dynamics in the reaction catalyzed by dihydrofolate
reductase from the hyperthermophile Thermotoga maritima (TmDHFR) has been examined by enzyme isotope substitution (<sup>15</sup>N, <sup>13</sup>C, <sup>2</sup>H). In contrast to all other
enzyme reactions investigated previously, including DHFR from Escherichia coli (EcDHFR), for which isotopic substitution
led to decreased reactivity, the rate constant for the hydride transfer
step is not affected by isotopic substitution of TmDHFR. TmDHFR therefore
appears to lack the coupling of protein motions to the reaction coordinate
that have been identified for EcDHFR catalysis. Clearly, dynamical
coupling is not a universal phenomenon that affects the efficiency
of enzyme catalysis