Structural and Spectroscopic Investigations of pH-Dependent Mo(V) Species in a Bacterial Sulfite-Oxidizing Enzyme.

Mono-pyranopterin-containing sulfite-oxidizing enzymes (SOEs), including eukaryotic sulfite oxidases and homologous prokaryotic sulfite dehydrogenases (SDHs), are molybdenum enzymes that exist in almost all forms of life, where they catalyze the direct oxidation of sulfite into sulfate, playing a key role in protecting cells and organisms against sulfite-induced damage. To decipher their catalytic mechanism, we have previously provided structural and spectroscopic evidence for direct coordination of HPO to the Mo atom at the active site of the SDH from the hyperthermophilic bacterium (SDH), mimicking the proposed sulfate-bound intermediate proposed to be formed during catalysis. In this work, by solving the X-ray crystallographic structure of the unbound enzyme, we resolve the changes in the hydrogen bonding network in the molybdenum environment that enable the stabilization of the previously characterized phosphate adduct.

Inverted All-Inorganic Nanorod-Based Light-Emitting Diodes via Electrophoretic Deposition.

High performance and high stability in all-inorganic solution processed nanocrystal-based light-emitting diodes (LEDs) are highly attractive for large area devices compared to organic material-based LEDs. In this work, an inverted all-inorganic LED structure is designed to have an easy integration with thin-film transistors. Adopting robust inorganic materials such as Ni O nanoparticle films as a hole transport layer (HTL) is beneficial for the performance of LED.

Contribution of a C-Terminal Extension to the Substrate Affinity and Oligomeric Stability of Aldehyde Dehydrogenase from HB27.

Aldehyde dehydrogenase enzymes (ALDHs) are widely studied for their roles in disease propagation and cell metabolism. Their use in biocatalysis applications, for the conversion of aldehydes to carboxylic acids, has also been recognized. Understanding the structural features and functions of both prokaryotic and eukaryotic ALDHs is key to uncovering novel applications of the enzyme and probing its role in disease propagation.

Individual heme a and heme a contributions to the Soret absorption spectrum of the reduced bovine cytochrome c oxidase.

Bovine cytochrome c oxidase (CcO) contains two hemes, a and a, chemically identical but differing in coordination and spin state. The Soret absorption band of reduced aa-type cytochrome c oxidase consists of overlapping bands of the hemes a and a. It shows a peak at ∼444 nm and a distinct shoulder at ∼425 nm.

Enzyme Immobilization on Metal Organic Frameworks: the Effect of Buffer on the Stability of the Support.

Metal organic frameworks (MOFs) have been used to encapsulate an array of enzymes in a rapid and facile manner; however, the stability of MOFs as supports for enzymes has not been examined in detail. This study examines the stability of MOFs with different compositions (Fe-BTC, Co-TMA, Ni-TMA, Cu-TMA, and ZIF-zni) in buffered solutions commonly used in enzyme immobilization and biocatalysis. Stability was assessed via quantification of the release of metals by inductively coupled plasma optical emission spectroscopy.

Long acting injectables for therapeutic proteins.

Biotherapeutic development presents a myriad of challenges in relation to delivery, in particular for protein therapeutics. Protein delivery is complicated due to hydrophilicity, size, rate of degradation in vivo, low permeation through biological barriers, pH and temperature sensitivity, as well as the need to conserve its quaternary structure to retain function. To preserve therapeutic levels in vivo, proteins require frequent administration due to their short half-lives.

Expression, Purification, and Enzyme Activity Assay of a Recombinant Aldehyde Dehydrogenase from , Using an Host.

Based on previous in-depth characterisation, aldehyde dehydrogenases (ALDH) are a diverse superfamily of enzymes, in terms of both structure and function, present in all kingdoms of life. They catalyse the oxidation of an aldehyde to carboxylic acid using the cofactor nicotinamide adenine dinucleotide (phosphate) (NAD(P)), and are often not substrate-specific, but rather have a broad range of associated biological functions, including detoxification and biosynthesis. We studied the structure of ALDH from , as well as performed its biochemical characterisation.

Histological Injury to Rat Brain, Liver, and Kidneys by Gold Nanoparticles is Dose-Dependent.

Gold nanoparticles (GNPs) possess various interesting plasmonic properties that can provide a variety of diagnostic and therapeutic functionalities for biomedical applications. Compared to other inorganic metal nanoparticles (NPs), GNPs are less toxic and more biocompatible. However, the toxicity of gold nanoparticles on humans can be significant due to the size effect.

Study of ALDH from -Expression, Purification and Characterisation of the Non-Substrate Specific, Thermophilic Enzyme Displaying Both Dehydrogenase and Esterase Activity.

Aldehyde dehydrogenases (ALDH), found in all kingdoms of life, form a superfamily of enzymes that primarily catalyse the oxidation of aldehydes to form carboxylic acid products, while utilising the cofactor NAD(P). Some superfamily members can also act as esterases using -nitrophenyl esters as substrates. The ALDH from was recombinantly expressed in and purified to obtain high yields (approximately 15-20 mg/L) and purity utilising an efficient heat treatment step coupled with IMAC and gel filtration chromatography.

Modulating the pro-apoptotic activity of cytochrome c at a biomimetic electrified interface.

Programmed cell death via apoptosis is a natural defence against excessive cell division, crucial for fetal development to maintenance of homeostasis and elimination of precancerous and senescent cells. Here, we demonstrate an electrified liquid biointerface that replicates the molecular machinery of the inner mitochondrial membrane at the onset of apoptosis. By mimicking in vivo cytochrome c (Cyt c) interactions with cell membranes, our platform allows us to modulate the conformational plasticity of the protein by simply varying the electrochemical environment at an aqueous-organic interface.

Balanced lipase interactions for degradation-controlled paclitaxel release from lipid cubic phase formulations.

Lipid cubic phase (LCP) formulations enhance the intestinal solubility and bioavailability of hydrophobic drugs by reducing precipitation and facilitating their mass transport to the intestinal surface for absorption. LCPs with an ester linkage connecting the acyl chain to the glycerol backbone (monoacylglycerols), are susceptible to chemical digestion by several lipolytic enzymes including lipases, accelerating the release of hydrophobic agents from the lipid bilayers of the matrix. Unlike regular enzymes that transform soluble substrates, lipolytic enzymes act at the interface of water and insoluble lipid.

Lipid Cubic Systems for Sustained and Controlled Delivery of Antihistamine Drugs.

Antihistamines are capable of blocking mediator responses in allergic reactions including allergic rhinitis and dermatological reactions. By incorporating various H receptor antagonists into a lipid cubic phase network, these active ingredients can be delivered locally over an extended period of time owing to the mucoadhesive nature of the system. Local delivery can avoid inducing unwanted side effects, often observed after systematic delivery.

Candidates for smart cardiovascular medical device coatings: A comparative study with endothelial and smooth muscle cells.

Stent-induced vascular injury is manifested by removal of the endothelium and phenotypic changes in the underlying medial smooth muscle cells layer. This results in pathological vascular remodelling primarily contributed to smooth muscle cell proliferation and leads to vessel re-narrowing; neointimal hyperplasia. Current drug-eluting stents release non-selective anti-proliferative drugs such as paclitaxel from the stent surface that not only inhibit growth of smooth muscle cells but also delay endothelial healing, potentially leading to stent thrombosis.

Stent conditioned media for in vitro evaluation of hydrophobic stent coatings.

In vitro cell studies of hydrophobic drugs face difficulties associated with their low aqueous solubility. To study poorly soluble drugs in bio-relevant media, solubilizing agents are frequently used to make stock solutions before final reconstitution in media. This results in drug concentrations that are not representative of in vivo conditions and may pose adverse effects on cells' biological functions.

Insights into Aldehyde Dehydrogenase Enzymes: A Structural Perspective.

Aldehyde dehydrogenases engage in many cellular functions, however their dysfunction resulting in accumulation of their substrates can be cytotoxic. ALDHs are responsible for the NAD(P)-dependent oxidation of aldehydes to carboxylic acids, participating in detoxification, biosynthesis, antioxidant and regulatory functions. Severe diseases, including alcohol intolerance, cancer, cardiovascular and neurological diseases, were linked to dysfunctional ALDH enzymes, relating back to key enzyme structure.

Specific inhibition of proton pumping by the T315V mutation in the K channel of cytochrome ba from Thermus thermophilus.

Cytochrome ba from Thermus thermophilus belongs to the B family of heme-copper oxidases and pumps protons across the membrane with an as yet unknown mechanism. The K channel of the A family heme-copper oxidases provides delivery of a substrate proton from the internal water phase to the binuclear heme-copper center (BNC) during the reductive phase of the catalytic cycle, while the D channel is responsible for transferring both substrate and pumped protons. By contrast, in the B family oxidases there is no D-channel and the structural equivalent of the K channel seems to be responsible for the transfer of both categories of protons.

Dark Field and Coherent Anti-Stokes Raman (DF-CARS) Imaging of Cell Uptake of Core-Shell, Magnetic-Plasmonic Nanoparticles.

Magnetic-plasmonic, FeO-Au, core-shell nanoparticles are popular in many applications, most notably in therapeutics and diagnostics, and thus, the imaging of these nanostructures in biological samples is of high importance. These nanostructures are typically imaged in biological material by dark field scatter imaging, which requires an even distribution of nanostructures in the sample and, therefore, high nanoparticle doses, potentially leading to toxicology issues. Herein, we explore the nonlinear optical properties of magnetic nanoparticles coated with various thicknesses of gold using the open aperture z-scan technique to determine the nonlinear optical properties and moreover, predict the efficacy of the nanostructures in nonlinear imaging.

Citric acid functionalized nitinol stent surface promotes endothelial cell healing.

While drug-eluting stents containing anti-proliferative agents inhibit proliferation of smooth muscle cells (SMCs), they also delay the regrowth of the endothelial cells which can result in subsequent development of restenosis. Acidic extracellular environments promote cell anchorage and migration by inducing conformational change in integrins, the main cell adhesion proteins. This study addresses the feasibility of a citric acid (CA) functionalized nitinol stent for improving vascular biocompatibility, specifically enhancing endothelialization.

Structural evidence for a reaction intermediate mimic in the active site of a sulfite dehydrogenase.

By combining X-ray crystallography, electron paramagnetic resonance techniques and density functional theory-based modelling, we provide evidence for a direct coordination of the product analogue, phosphate, to the molybdenum active site of a sulfite dehydrogenase. This interaction is mimicking the still experimentally uncharacterized reaction intermediate proposed to arise during the catalytic cycle of this class of enzymes. This work opens new perspectives for further deciphering the reaction mechanism of this nearly ubiquitous class of oxidoreductases.

Modulating the release of pharmaceuticals from lipid cubic phases using a lipase inhibitor.

Lipid cubic phase formulations have gained recognition as potential controlled delivery systems for a range of active pharmaceutical and biological agents on account of their desirable physiochemical properties and ability to encapsulate both hydrophobic and hydrophilic molecules. The most widely studied lipid cubic systems are those of the monoacylglycerol lipid family. These formulations are susceptible to lipolysis by a variety of enzymes, including lipases and esterases, which attack the ester bond present on the lipid chain bridging the oleic acid component to the glycerol backbone.

The Reactions of O and NO with Mixed-Valence ba Cytochrome c Oxidase from Thermus thermophilus.

Earlier CO flow-flash experiments on the fully reduced Thermus thermophilus ba (Tt ba) cytochrome oxidase revealed that O binding was slowed down by a factor of 10 in the presence of CO (Szundi et al., 2010, PNAS 107, 21010-21015). The goal of the current study is to explore whether the long apparent lifetime (∼50 ms) of the Cu-CO complex generated upon photolysis of the CO-bound mixed-valence Tt ba (Koutsoupakis et al.

Pyruvate Decarboxylase is Less Effective Than That of for Ethanol Production in Metabolically Engineered sp. PCC6803.

To produce bioethanol from model cyanobacteria such as , a two gene cassette consisting of genes encoding pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH) are required to transform pyruvate first to acetaldehyde and then to ethanol. However the partition of pyruvate to ethanol comes at a cost, a reduction in biomass and pyruvate availability for other metabolic processes. Hence strategies to divert flux to ethanol as a biofuel in are of interest.

APTES Duality and Nanopore Seed Regulation in Homogeneous and Nanoscale-Controlled Reduction of Ag Shell on SiO Microparticle for Quantifiable Single Particle SERS.

Noble-metal nanoparticles size and packing density are critical for sensitive surface-enhanced Raman scattering (SERS) and controlled preparation of such films required to achieve reproducibility. Provided that they are made reliable, Ag shell on SiO microscopic particles (Ag/SiO) are promising candidates for lab-on-a-bead analytical measurements of low analyte concentration in liquid specimen. Here, we selected nanoporous silica microparticles as a substrate for reduction of AgNO with 3-aminopropyltriethoxysilane (APTES).

Integral caa-Cytochrome c Oxidase from Thermus thermophilus: Purification and Crystallization.

Cytochrome c oxidase is a respiratory enzyme catalyzing the energy-conserving reduction of molecular oxygen to water-a fundamental biological process of cell respiration. The first crystal structures of the type A cytochrome c oxidases, bovine heart and Paracoccus denitrificans cytochrome c oxidases, were published in 1995 and contributed immensely to the understanding of the enzyme's mechanism of action. The senior author's research focus was directed toward understanding the structure and function of the type B cytochrome c oxidases, ba-oxidase and type A2 caa-oxidase, both from the extreme thermophilic bacterium Thermus thermophilus.