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.

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.