Quantification of guanidine in environmental samples using benzoin derivatization and LC-MS analysis.

The recent discovery of guanidine-dependent riboswitches in many microbes raised interest in the biological function and metabolism of this nitrogen-rich compound. However, very little is known about the concentrations of guanidine in the environment. Several methods have been published for quantifying guanidine and guanidino compounds in human urine and blood, often relying on derivatization followed by fluorescence detection.

Growth of complete ammonia oxidizers on guanidine.

Guanidine is a chemically stable nitrogen compound that is excreted in human urine and is widely used in manufacturing of plastics, as a flame retardant and as a component of propellants, and is well known as a protein denaturant in biochemistry. Guanidine occurs widely in nature and is used by several microorganisms as a nitrogen source, but microorganisms growing on guanidine as the only substrate have not yet been identified. Here we show that the complete ammonia oxidizer (comammox) Nitrospira inopinata and probably most other comammox microorganisms can grow on guanidine as the sole source of energy, reductant and nitrogen.

Biodegradation of Water-Soluble Polymers by Wastewater Microorganisms: Challenging Laboratory Testing Protocols.

For water-soluble polymers (WSPs) that enter environmental systems at their end-of-life, biodegradability is a key functionality. For the development and regulation of biodegradable WSPs, testing methods that are both scientifically validated and economically practicable are needed. Here, we used respirometric laboratory tests to study the biodegradation of poly(amino acids), poly(ethylene glycol), and poly(vinyl alcohol), together with appropriate low-molecular-weight reference substrates.

Hydrolysis of Antimicrobial Peptides by Extracellular Peptidases in Wastewater.

Several antimicrobial peptides (AMPs) are emerging as promising novel antibiotics. When released into wastewater streams after use, AMPs might be hydrolyzed and inactivated by wastewater peptidases─resulting in a reduced release of active antimicrobials into wastewater-receiving environments. A key step towards a better understanding of the fate of AMPs in wastewater systems is to investigate the activity and specificity of wastewater peptidases.

Uptake, Metabolism and Accumulation of Tire Wear Particle-Derived Compounds in Lettuce.

Tire wear particle (TWP)-derived compounds may be of high concern to consumers when released in the root zone of edible plants. We exposed lettuce plants to the TWP-derived compounds diphenylguanidine (DPG), hexamethoxymethylmelamine (HMMM), benzothiazole (BTZ), -phenyl-N'-(1,3-dimethylbutyl)--phenylenediamine (6PPD), and its quinone transformation product (6PPD-q) at concentrations of 1 mg L in hydroponic solutions over 14 days to analyze if they are taken up and metabolized by the plants. Assuming that TWP may be a long-term source of TWP-derived compounds to plants, we further investigated the effect of leaching from TWP on the concentration of leachate compounds in lettuce leaves by adding constantly leaching TWP to the hydroponic solutions.

Effect of Polymer Properties on the Biodegradation of Polyurethane Microplastics.

The release of fragments from plastic products, that is, secondary microplastics, is a major concern in the context of the global plastic pollution. Currently available (thermoplastic) polyurethanes [(T)PU] are not biodegradable and therefore should be recycled. However, the ester bond in (T)PUs might be sufficiently hydrolysable to enable at least partial biodegradation of polyurethane particles.

Environmental Biodegradation of Water-Soluble Polymers: Key Considerations and Ways Forward.

Water-soluble polymers (WSPs) have unique properties that are valuable in diverse applications ranging from home and personal care products to agricultural formulations. For applications that result in the release of WSPs into natural environments or engineered systems, such as agricultural soils and wastewater streams, biodegradable as opposed to nonbiodegradable WSPs have the advantage of breaking down and, thereby, eliminating the risk of persistence and accumulation. In this Commentary, we emphasize central steps in WSP biodegradation, discuss how these steps depend on both WSP properties and characteristics of the receiving environment, and highlight critical requirements for testing WSP biodegradability.

Identifying Functional Groups that Determine Rates of Micropollutant Biotransformations Performed by Wastewater Microbial Communities.

The goal of this research was to identify functional groups that determine rates of micropollutant (MP) biotransformations performed by wastewater microbial communities. To meet this goal, we performed a series of incubation experiments seeded with four independent wastewater microbial communities and spiked them with a mixture of 40 structurally diverse MPs. We collected samples over time and used high-resolution mass spectrometry to estimate biotransformation rate constants for each MP in each experiment and to propose structures of 46 biotransformation products.

Towards more Sustainable Peptide- based Antibiotics: Stable in Human Blood, Enzymatically Hydrolyzed in Wastewater?

The emergence and spread of antibiotic resistance is a major societal challenge and new antibiotics are needed to successfully fight bacterial infections. Because the release of antibiotics into wastewater and downstream environments is expected to contribute to the problem of antibiotic resistance, it would be beneficial to consider the environmental fate of antibiotics in the development of novel antibiotics. In this article, we discuss the possibility of designing peptide-based antibiotics that are stable during treatment ( in human blood), but rapidly inactivated through hydrolysis by peptidases after their secretion into wastewater.

Exploring the Specificity of Extracellular Wastewater Peptidases to Improve the Design of Sustainable Peptide-Based Antibiotics.

New antimicrobial peptides are emerging as promising alternatives to conventional antibiotics because of their specificity for target pathogens and their potential to be rapidly hydrolyzed (, inactivated) by extracellular peptidases during biological wastewater treatment, thereby limiting the emergence and propagation of antibiotic resistance in the environment. However, little is known about the specificity of extracellular peptidases derived from wastewater microbial communities, which is a major impediment for the design of sustainable peptide-based antibiotics that can be hydrolyzed by wastewater peptidases. We used a set of natural peptides to explore the specificity of dissolved extracellular wastewater peptidases.

Biotransformation of antibiotics: Exploring the activity of extracellular and intracellular enzymes derived from wastewater microbial communities.

Evaluating the activity of extracellular and intracellular enzymes derived from wastewater microbial communities is essential to improve our fundamental understanding of micropollutant removal during wastewater treatment. To study biotransformations with respect to enzyme biogeography, we developed a method to separate soluble extracellular, extracellular polymeric substance (EPS)-bound, and intracellular enzymes from wastewater microbial communities and assessed the protease and peptidase activity of the resulting enzyme pools. We also evaluated the biotransformation of six antibiotics (amoxicillin, ampicillin, clindamycin, daptomycin, linezolid, and vancomycin) in each enzyme pool because we expect that the kinetics, pathways, and biogeography of antibiotic biotransformations influence the selection of antibiotic resistance within wastewater microbial communities and in downstream environments.

Photochemical Transformation of Poly(butylene adipate- co-terephthalate) and Its Effects on Enzymatic Hydrolyzability.

Biodegradable polyesters are being increasingly used to replace conventional, nondegradable polymers in agricultural applications such as plastic film for mulching. For many of these applications, poly(butylene adipate- co-terephthalate) (PBAT) is a promising biodegradable material. However, PBAT is also susceptible to photochemical transformations.

Sustainable Polyester Elastomers from Lactones: Synthesis, Properties, and Enzymatic Hydrolyzability.

Chemically cross-linked elastomers are an important class of polymeric materials with excellent temperature and solvent resistance. However, nearly all elastomers are petroleum-derived and persist in the environment or in landfills long after they are discarded; this work strives to address these issues by demonstrating the synthesis of renewable, enzymatically hydrolyzable, and mechanically competitive polyester elastomers. The elastomers described were synthesized using a novel bis(β-lactone) cross-linker and star-shaped, hydroxyl-terminated poly(γ-methyl-ε-caprolactone).

Polyol Structure Influences Enzymatic Hydrolysis of Bio-Based 2,5-Furandicarboxylic Acid (FDCA) Polyesters.

Polyesters of 2,5-furandicarboxylic acid (FDCA) have gained attention as they can be regarded as the bio-based alternatives to the petroleum-based polyesters of terephthalic acid. However, only little is known about the biodegradation and enzymatic hydrolysis of FDCA-based polyesters. This work aims to investigate the influence of different polyols on enzymatic hydrolysis of FDCA-based polyesters.

Enzymatic Degradation of Aromatic and Aliphatic Polyesters by Expressed Cutinase 1 from .

To study hydrolysis of aromatic and aliphatic polyesters cutinase 1 from (Thc_Cut1) was expressed in . No significant differences between the expression of native Thc_Cut1 and of two glycosylation site knock out mutants (Thc_Cut1_koAsn and Thc_Cut1_koST) concerning the total extracellular protein concentration and volumetric activity were observed. Hydrolysis of poly(ethylene terephthalate) (PET) was shown for all three enzymes based on quantification of released products by HPLC and similar concentrations of released terephthalic acid (TPA) and mono(2-hydroxyethyl) terephthalate (MHET) were detected for all enzymes.

Enzymatic Hydrolysis of Polyester Thin Films at the Nanoscale: Effects of Polyester Structure and Enzyme Active-Site Accessibility.

Biodegradable polyesters have a large potential to replace persistent polymers in numerous applications and to thereby reduce the accumulation of plastics in the environment. Ester hydrolysis by extracellular carboxylesterases is considered the rate-limiting step in polyester biodegradation. In this work, we systematically investigated the effects of polyester and carboxylesterase structure on the hydrolysis of nanometer-thin polyester films using a quartz-crystal microbalance with dissipation monitoring.

High-Throughput Analysis of Enzymatic Hydrolysis of Biodegradable Polyesters by Monitoring Cohydrolysis of a Polyester-Embedded Fluorogenic Probe.

Biodegradable polyesters have the potential to replace nondegradable, persistent polymers in numerous applications and thereby alleviate plastic accumulation in the environment. Herein, we present an analytical approach to study enzymatic hydrolysis of polyesters, the key step in their overall biodegradation process. The approach is based on embedding fluorescein dilaurate (FDL), a fluorogenic ester substrate, into the polyester matrix and on monitoring the enzymatic cohydrolysis of FDL to fluorescein during enzymatic hydrolysis of the polyester.

Uukuniemi Virus as a Tick-Borne Virus Model.

Unlabelled: In the last decade, novel tick-borne pathogenic phleboviruses in the family Bunyaviridae, all closely related to Uukuniemi virus (UUKV), have emerged on different continents. To reproduce the tick-mammal switch in vitro, we first established a reverse genetics system to rescue UUKV with a genome close to that of the authentic virus isolated from the Ixodes ricinus tick reservoir. The IRE/CTVM19 and IRE/CTVM20 cell lines, both derived from I.

Enzymatic Hydrolysis of Polyester Thin Films: Real-Time Analysis of Film Mass Changes and Dissipation Dynamics.

Cleavage of ester bonds by extracellular microbial hydrolases is considered a key step during the breakdown of biodegradable polyester materials in natural and engineered systems. Here we present a novel analytical approach for simultaneous detection of changes in the masses and rigidities of polyester thin films during enzymatic hydrolysis using a Quartz Crystal Microbalance with Dissipation monitoring (QCM-D). In experiments with poly(butylene succinate) (PBS) and the lipase of Rhizopus oryzae (RoL), we detected complete hydrolysis of PBS thin films at pH 5 and 40 °C that proceeded through soft and water-rich film intermediates.

Biomimetic Approach to Enhance Enzymatic Hydrolysis of the Synthetic Polyester Poly(1,4-butylene adipate): Fusing Binding Modules to Esterases.

Mimicking a concept of nature for the hydrolysis of biopolymers, the Thermobifida cellulosilytica cutinase 1 (Thc_Cut1) was fused to a polymer binding module (PBM) to enhance the hydrolysis of the polyester poly(1,4-butylene adipate) (PBA). Namely, the binding module of a polyhydroxyalkanoate depolymerase from Alcaligenes faecalis (Thc_Cut1_PBM) was attached to the cutinase via two different linker sequences varying in length. In order to investigate the adsorption behavior, catalytically inactive mutants both of Thc_Cut1 and Thc_Cut1_PBM were successfully constructed by site-directed mutagenesis of serine 131 to alanine.

Host cell entry of respiratory syncytial virus involves macropinocytosis followed by proteolytic activation of the F protein.

Respiratory Syncytial Virus (RSV) is a highly pathogenic member of the Paramyxoviridae that causes severe respiratory tract infections. Reports in the literature have indicated that to infect cells the incoming viruses either fuse their envelope directly with the plasma membrane or exploit clathrin-mediated endocytosis. To study the entry process in human tissue culture cells (HeLa, A549), we used fluorescence microscopy and developed quantitative, FACS-based assays to follow virus binding to cells, endocytosis, intracellular trafficking, membrane fusion, and infection.