β2-Microglobulin Amyloid Fibrils Are Nanoparticles That Disrupt Lysosomal Membrane Protein Trafficking and Inhibit Protein Degradation by Lysosomes

Fragmentation of amyloid fibrils produces fibrils that are reduced in length but have an otherwise unchanged molecular architecture. The resultant nanoscale fibril particles inhibit the cellular reduction of the tetrazolium dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), a substrate commonly used to measure cell viability, to a greater extent than unfragmented fibrils. Here we show that the internalization of β₂-microglobulin (β₂m) amyloid fibrils is dependent on fibril length, with fragmented fibrils being more efficiently internalized by cells. Correspondingly, inhibiting the internalization of fragmented β₂m fibrils rescued cellular MTT reduction. Incubation of cells with fragmented β₂m fibrils did not, however, cause cell death. Instead, fragmented β₂m fibrils accumulate in lysosomes, alter the trafficking of lysosomal membrane proteins, and inhibit the degradation of a model protein substrate by lysosomes. These findings suggest that nanoscale fibrils formed early during amyloid assembly reactions or by the fragmentation of longer fibrils could play a role in amyloid disease by disrupting protein degradation by lysosomes and trafficking in the endolysosomal pathway.     

Biodegradation of polyurethane: a review

Lack of degradability and the closing of landfill sites as well as growing water and land pollution problems have led to concern about plastics. Increasingly, raw materials such as crude oil are in short supply for the synthesis of plastics, and the recycling of waste plastics is becoming more important. As the importance of recycling increases, so do studies on elucidation of the biodegradability of polyurethanes. Polyurethanes are an important and versatile class of man-made polymers used in a wide variety of products in the medical, automotive and industrial fields. Polyurethane is a general term used for a class of polymers derived from the condensation of polyisocyanates and polyalcohols. Despite its xenobiotic origins, polyurethane has been found to be susceptible to biodegradation by naturally occurring microorganisms. Microbial degradation of polyurethanes is dependent on the many properties of the polymer such as molecular orientation, crystallinity, cross-linking and chemical groups present in the molecular chains which determine the accessibility to degrading-enzyme systems. Esterase activity (both membrane-bound and extracellular) has been noted in microbes which allow them to utilize polyurethane. Microbial degradation of polyester polyurethane is hypothosized to be mainly due to the hydrolysis of ester bonds by these esterase enzymes.     

Use of LCA indicators to assess Iranian rapeseed production systems with different residue management practices

In this study the environmental profile of Iranian rapeseed cultivation was analysed with a Life Cycle Assessment (LCA) approach, in order to identify the hot spots of the system. Also, in order to apply environmental indicators for agricultural decision making purposes, the environmental burden of different residue management practices were compared. Primary data were collected from 150 rapeseed farms from the Iranian Mazandaran province. The system considers a cradle-to-farm gate boundary, and the functional unit was regarded as being one Mg of rapeseed production. The LCA results indicate that the global warming potential amounts to 1181.6 kg CO_2eq Mg⁻¹. Also, the acidification and eutrophication potentials were found to be 23.3 kg SO_2eq Mg⁻¹ and 18 kg PO₄³_eq Mg⁻¹, respectively. The results also reveal that environmental emissions of crop production were significantly affected by residue management practices. Specifically, rapeseed residue removal from the field is the most environmentally-friendly practice. This is followed by a scenario involving residue incorporation in the soil. The practice of burning such residue entails the highest environmental emissions. In conclusion, reducing the consumption of chemical fertilisers, especially nitrogen based ones, is important for decreasing the environmental footprints in the area. Furthermore, avoiding crop residue burning and developing rapeseed-bean rotation are favourable management strategies for establishing more environmentally-friendly rapeseed production systems in the region.     

A pH-stable laccase from alkali-tolerant γ-proteobacterium JB: Purification, characterization and indigo carmine degradation

γ-Proteobacterium JB, an alkali-tolerant soil isolate, produced laccase constitutively in unbuffered medium. The enzyme was purified to homogeneity by ammonium sulphate precipitation, DEAE-sepharose anion exchange chromatography and preparatory polyacrylamide gel electrophoresis. The purified enzyme was a monomeric polypeptide (MW 120 kDa) and absorbed at 590 nm indicating the presence of Type I Cu²⁺-centre. It worked optimally at 55 °C and showed different pH optima for different substrates. The enzyme was highly stable in the pH range 4–10 even after 60 days at 4 °C. K_m and V_max values for syringaldazine, catechol, pyrogallol, p-phenylenediamine, l-methyl DOPA and guaiacol substrates were determined. Inhibitors, viz. azide, diethyldithiocarbamate, thioglycollate and cysteine-hydrochloride all inhibited laccase non-competitively using guaiacol as substrate at pH 6.5. The enzyme degraded indigo carmine (pH 9, 55 °C) to anthranilic acid via isatin as determined spectrophotometrically and by HPLC analysis. Degradation was enhanced in the presence of syringaldehyde (571%), vanillin (156%) and p-hydroxybenzoic acid (91.6%) but not HOBT.     

A Golgi-localized Mannosidase (MAN1B1) Plays a Non-enzymatic Gatekeeper Role in Protein Biosynthetic Quality Control

Conformation-based disorders are manifested at the level of protein structure, necessitating an accurate understanding of how misfolded proteins are processed by the cellular proteostasis network. Asparagine-linked glycosylation plays important roles for protein quality control within the secretory pathway. The suspected role for the MAN1B1 gene product MAN1B1, also known as ER mannosidase I, is to function within the ER similar to the yeast ortholog Mns1p, which removes a terminal mannose unit to initiate a glycan-based ER-associated degradation (ERAD) signal. However, we recently discovered that MAN1B1 localizes to the Golgi complex in human cells and uncovered its participation in ERAD substrate retention, retrieval to the ER, and subsequent degradation from this organelle. The objective of the current study was to further characterize the contribution of MAN1B1 as part of a Golgi-based quality control network. Multiple lines of experimental evidence support a model in which neither the mannosidase activity nor catalytic domain is essential for the retention or degradation of the misfolded ERAD substrate Null Hong Kong. Instead, a highly conserved, vertebrate-specific non-enzymatic decapeptide sequence in the luminal stem domain plays a significant role in controlling the fate of overexpressed Null Hong Kong. Together, these findings define a new functional paradigm in which Golgi-localized MAN1B1 can play a mannosidase-independent gatekeeper role in the proteostasis network of higher eukaryotes.     

Enantioselective toxicity and degradation of chiral herbicide fenoxaprop-ethyl in earthworm Eisenia fetida

The enantioselective degradation of fenoxaprop-ethyl in ecological indicator earthworm was studied and the main metabolites (fenoxaprop, 6-chloro-2,3-dihydrobenzoxazol-2-one, ethyl-2-(4-hydroxyphenoxy)propanoate, 2-(4-hydroxyphenoxy)propanoic acid) were also monitored on an enantiomeric level. The individual enantiomers of fenoxaprop-ethyl and its three chiral metabolites were prepared to study the acute toxicity to earthworm. Chiral analysis methods were set up based on HPLC–MS/MS with chiralpak IC chiral column. Fenoxaprop-ethyl was not found in earthworms, while the primary metabolite fenoxaprop was in relatively high levels indicating a quick hydrolysis degradation. Fenoxaprop was accumulated almost exclusively with R-enantiomer in earthworms and the bio-concentration factors of R-fenoxaprop and S-fenoxaprop were 1.39 and 0.17 respectively with the enantiomer fraction (EF) values about 0.99. The degradation of R-fenoxaprop in earthworms followed first-order kinetics with half-life of 1.82 day. The other metabolites could not be detected in earthworms. The calculated LC₅₀ values showed ecological indicator earthworm was more sensitive to the four metabolites than fenoxaprop-ethyl. Furthermore, earthworm was more sensitive to the R-form of the chiral metabolites than the S-form and rac-form. The results suggested metabolites and enantioselectivity should be taken into consideration to better predict the exposure concentration and apply ecological indicators in toxicological studies.     

Pharmacokinetic studies of naproxen amides of some amino acid esters with promising colorectal cancer chemopreventive activity

Naproxen (nap) is belonging to Non-steriodal anti-inflammatory drugs (NSAIDs) group of drugs that characterized by their free carboxylic group. The therapeutic activity of nap is usually accompanied by GI untoward side effects. Recently synthesized naproxen amides of some amino acid esters prodrugs to mask the free carboxylic group were reported. Those prodrugs showed a promising colorectal cancer chemopreventive activity. The current study aims to investigate the fate and hydrolysis of the prodrugs kinetically in different pH conditions, simulated gastric and intestinal fluids with pHs of 1.2, 5.5 and 7.4 in vitro at 37 °C. The effect of enzymes on the hydrolysis of prodrugs was also studied through incubation of these prodrugs at 37 °C in human plasma and rat liver homogenates. The pharmacokinetic parameters of selected prodrugs and the liberated nap were studied after oral and intraperitoneal administration in male wistar rats. The results showed the hydrolysis of naproxen amides of amino acid esters to nap through two steps first by degradation of the ester moiety to form the amide of nap with amino acid and the second was through the degradation of the amide link to liberate nap. The two reactions were followed and studied kinetically where K₁ and K₂ (rate constants of degradation) is reported. The hydrolysis of prodrugs was faster in liver homogenates than in plasma. The relative bioavailability of the liberated nap in vivo was higher in case of prodrug containing ethyl glycinate moiety than that occupied l-valine ethyl ester moiety. Each of nap. prodrugs containing ethyl glycinate and l-valine ethyl ester moieties appears promising in liberating nap, decreasing direct GI side effect and consequently their colorectal cancer chemopreventive activity.     

NADH Binds and Stabilizes the 26S Proteasomes Independent of ATP

The 26S proteasome is the end point of the ubiquitin- and ATP-dependent degradation pathway. The 26S proteasome complex (26S PC) integrity and function has been shown to be highly dependent on ATP and its homolog nucleotides. We report here that the redox molecule NADH binds the 26S PC and is sufficient in maintaining 26S PC integrity even in the absence of ATP. Five of the 19S proteasome complex subunits contain a putative NADH binding motif (GxGxxG) including the AAA-ATPase subunit, Psmc1 (Rpt2). We demonstrate that recombinant Psmc1 binds NADH via the GxGxxG motif. Introducing the ΔGxGxxG Psmc1 mutant into cells results in reduced NADH-stabilized 26S proteasomes and decreased viability following redox stress induced by the mitochondrial inhibitor rotenone. The newly identified NADH binding of 26S proteasomes advances our understanding of the molecular mechanisms of protein degradation and highlights a new link between protein homeostasis and the cellular metabolic/redox state.     

Biodegradation of octyltin compounds by Cochliobolus lunatus and influence of xenobiotics on fungal fatty acid composition

The perturbation in the growth and fatty acid profile of the microscopic fungus Cochliobolus lunatus IM 4417 in the presence of octyltin compounds (trioctyltin – TOT, dioctyltin – DOT and monooctyltin – MOT) was studied. Fungal resistance to the tested organotins decreased with a reduction in the number of octyl groups bonded with a tin atom. Also, the fatty acid unsaturation index decreased according to the mentioned scheme. Among all tested octyltin compounds, TOT was removed with the highest efficiency. The efficiency of MOT removal was correlated with the initial concentration of the compounds and for concentrations 20 and 100 mg l⁻¹ reached the value of 75% and 40%, respectively. Elimination of octyltins depended on the metabolic activity of the fungus and was not the result of passive sorption. During bioconversion of TOT the hydroxylated derivative of substrate was detected. Moreover, the addition of cytochrome P-450 inhibitors significantly reduced the metabolism of octyltin compounds. Thus, it is postulated that the process of degradation of octyltin compounds is similar to that described for tributyltin (TBT) and it is mediated by cytochrome P-450.