Parameter Reliability and Understanding Enzyme Function.

Knowledge of the Michaelis-Menten parameters and their meaning in different circumstances is an essential prerequisite to understanding enzyme function and behaviour. The published literature contains an abundance of values reported for many enzymes. The problem concerns assessing the appropriateness and validity of such material for the purpose to which it is to be applied.

Enzyme nomenclature and classification: the state of the art.

The IUBMB enzyme classification system, available at the IUBMB ExplorEnz website, uses a four-component number (the EC number) that identifies an enzyme in terms of reaction catalysed. There were originally six recognized groups of enzymes: Oxidoreductases (EC 1), Transferases (EC 2), Hydrolases (EC 3), Lyases (EC 4), Isomerases (EC 5) and Ligases (EC 6). Of these, the lyases, which are defined as 'enzymes that cleave C-C, C-O, C-N and other bonds by means other than by hydrolysis or oxidation', present particular recognition and classification problems.

Degradation of thymic humoral factor γ2 in human, rat and mouse blood: An experimental and theoretical study.

The degradation of the immunomodulatory octapeptide, thymic humoral factor γ2 (THF-γ2, thymoctonan) has been studied in whole blood samples from human, rat and mouse. The peptide, Leu-Glu-Asp-Gly-Pro-Lys-Phe-Leu, was shown to be rapidly degraded by peptidases. The half-life of the intact peptide was less than 6 min at 37 °C in blood from the three species tested.

6-Hydroxydopamine: a far from simple neurotoxin.

6-Hydroxydopamine (6-OHDA), which is a neurotoxin that selectively destroys catecholaminergic nerves in sympathetically innervated tissues, has been used to provide a model of Parkinson's disease in experimental animals. It is rapidly autoxidised to yield potentially toxic products and reactive oxygen species. Its ability to release Fe(II) from protein storage sites also results in the formation of hROS.

The application of bathophenanthroline for the determination of free iron in parallel with hROS in microdialysis samples.

Background: Free, non-protein bound, Fe(II), which can catalyse the formation of the toxic highly-reactive oxygen species (hROS), has been implicated in several neurodegenerative conditions. The determination of free Fe(II) and Fe(III) in samples obtained from microdialysis experiments has been limited by the small amounts of sample available.

New Method: This work describes the development of a HPLC, with absorbance detection, method, based on the complexation of Fe(II) with bathophenanthroline disulfonate (BS), which allows a complete extracellular iron analysis with the small sample amounts that are available from in vivo microdialysis in rat brain.

Theobromine and related methylxanthines as inhibitors of Primary Amine Oxidase.

Methylxanthines are among the most widely consumed drugs in the world and evidence of their health benefits has been growing in recent years. Primary Amine Oxidase (PrAO) has been recognized as a therapeutic target for the amelioration of inflammatory, vascular, and neurodegenerative diseases. Previous work in our laboratories showed that caffeine inhibited Bovine PrAO with a Ki of 1.

A mechanism for bistability in glycosylation.

Glycosyltransferases are a class of enzymes that catalyse the posttranslational modification of proteins to produce a large number of glycoconjugate acceptors from a limited number of nucleotide-sugar donors. The products of one glycosyltransferase can be the substrates of several other enzymes, causing a combinatorial explosion in the number of possible glycan products. The kinetic behaviour of systems where multiple acceptor substrates compete for a single enzyme is presented, and the case in which high concentrations of an acceptor substrate are inhibitory as a result of abortive complex formation, is shown to result in non-Michaelian kinetics that can lead to bistability in an open system.

90 years of monoamine oxidase: some progress and some confusion.

It would not be practical to attempt to deal with all the advances that have informed our understanding of the behavior and functions of this enzyme over the past 90 years. This account concentrates key advances that explain why the monoamine oxidases remain of pharmacological and biochemical interest and on some areas of continuing uncertainty. Some issues that remain to be understood or are in need of further clarification are highlighted.

STRENDA DB: enabling the validation and sharing of enzyme kinetics data.

Standards for reporting enzymology data (STRENDA) DB is a validation and storage system for enzyme function data that incorporates the STRENDA Guidelines. It provides authors who are preparing a manuscript with a user-friendly, web-based service that checks automatically enzymology data sets entered in the submission form that they are complete and valid before they are submitted as part of a publication to a journal.

Assessment of Enzyme Inhibition: A Review with Examples from the Development of Monoamine Oxidase and Cholinesterase Inhibitory Drugs.

The actions of many drugs involve enzyme inhibition. This is exemplified by the inhibitors of monoamine oxidases (MAO) and the cholinsterases (ChE) that have been used for several pharmacological purposes. This review describes key principles and approaches for the reliable determination of enzyme activities and inhibition as well as some of the methods that are in current use for such studies with these two enzymes.

Monoaminergic and Histaminergic Strategies and Treatments in Brain Diseases.

The monoaminergic systems are the target of several drugs for the treatment of mood, motor and cognitive disorders as well as neurological conditions. In most cases, advances have occurred through serendipity, except for Parkinson's disease where the pathophysiology led almost immediately to the introduction of dopamine restoring agents. Extensive neuropharmacological studies first showed that the primary target of antipsychotics, antidepressants, and anxiolytic drugs were specific components of the monoaminergic systems.

Reflections on 60 years of publication of the Journal of Neurochemistry.

This review reflects on the origins, development, publishing trends, and scientific directions of the Journal of Neurochemistry over its 60 year lifespan as seen by key contributors to the Journal's production. The Journal first appeared in May 1956 with just two issues published in that inaugural year. By 1963, it appeared monthly and, by 2002, 24 hard copy issues were published yearly.

Multi-Target Directed Donepezil-Like Ligands for Alzheimer's Disease.

HIGHLIGHTS ASS234 is a MTDL compound containing a moiety from Donepezil and the propargyl group from the PF 9601N, a potent and selective MAO B inhibitor. This compound is the most advanced anti-Alzheimer agent for preclinical studies identified in our laboratory.Derived from ASS234 both multipotent donepezil-indolyl (MTDL-1) and donepezil-pyridyl hybrids (MTDL-2) were designed and evaluated as inhibitors of AChE/BuChE and both MAO isoforms.

A Knowledge-Based System for Display and Prediction of O-Glycosylation Network Behaviour in Response to Enzyme Knockouts.

O-linked glycosylation is an important post-translational modification of mucin-type protein, changes to which are important biomarkers of cancer. For this study of the enzymes of O-glycosylation, we developed a shorthand notation for representing GalNAc-linked oligosaccharides, a method for their graphical interpretation, and a pattern-matching algorithm that generates networks of enzyme-catalysed reactions. Software for generating glycans from the enzyme activities is presented, and is also available online.

Continuous monitoring of highly reactive oxygen radicals during in vivo microdialysis.

Background: Terephthalate (TA(2-)), which reacts with highly reactive oxygen species (hROS) to form the fluorophor 2-hydroxy terephthalic acid (OH-TA) with a high selectivity, has been used for determining hROS formation during in vivo microdialysis. Previously this involved collecting fractions of the microdialysate and determining the OH-TA formed after HPLC (the batch method).

New Method: This work reports the development and validation of a procedure for continuously determining hROS formation during microdialysis.

First-in-class thyrotropin-releasing hormone (TRH)-based compound binds to a pharmacologically distinct TRH receptor subtype in human brain and is effective in neurodegenerative models.

JAK4D, a first-in-class thyrotropin-releasing hormone (TRH)-based compound, is a prospective therapeutic candidate offering a multifaceted approach to treating neurodegeneration and other CNS conditions. The purpose of these studies was to determine the ability of JAK4D to bind to TRH receptors in human brain and to evaluate its neuropharmacological effects in neurodegenerative animal models. Additionally, JAK4D brain permeation was examined in mouse, and initial toxicology was assessed in vivo and in vitro.

Galactosyltransferase 4 is a major control point for glycan branching in N-linked glycosylation.

Protein N-glycosylation is a common post-translational modification that produces a complex array of branched glycan structures. The levels of branching, or antennarity, give rise to differential biological activities for single glycoproteins. However, the precise mechanism controlling the glycan branching and glycosylation network is unknown.

Neuroprotective effects of the MAO-B inhibitor, PF9601N, in an in vivo model of excitotoxicity.

Background: PF9601N [N-(2-propynyl)-2-(5-benzyloxy-indolyl) methylamine] is an inhibitor of monoamine oxidase B (MAO-B), which has shown to possess neuroprotective properties in several in vitro and in vivo models of Parkinson's disease (PD). As there is evidence that excitotoxicity may be implicated in the pathophysiology of several neurodegenerative diseases, the aim of the present work was to investigate the effects of PF9601N in an acute in vivo model of excitotoxicity induced by the local administration of kainic acid during striatal microdialysis in adult rats.

Methods: The basal and evoked release of neurotransmitters was monitored by HPLC analysis of microdialysate samples and tissue damage was evaluated histologically "ex vivo.

Effects of tyramine and 4-aminophenol on the oscillating peroxidase-oxidase reaction.

The peroxidase-oxidase oscillator, a model of biological oscillations, is usually studied in conjunction with the effector molecule, 2,4-dichlorophenol. In this account, we present evidence of the effects of a naturally occurring phenol, tyramine, on the reaction, and also those of the structurally similar 4-aminophenol. Whereas 2,4-dichlorophenol gives rise to sustained oscillations at 40 μM, it was discovered that tyramine promotes damped oscillations at a concentration of 120 μM.

Elucidation of metabolic pathways from enzyme classification data.

The IUBMB Enzyme List is widely used by other databases as a source for avoiding ambiguity in the recognition of enzymes as catalytic entities. However, it was not designed for metabolic pathway tracing, which has become increasingly important in systems biology. A Reactions Database has been created from the material in the Enzyme List to allow reactions to be searched by substrate/product, and pathways to be traced from any selected starting/seed substrate.

Fifty-five years of enzyme classification: advances and difficulties.

Since the publication of a list of enzymes classified according to the reactions that they catalysed, by Dixon and Webb in 1958, its content and presentation have undergone a number of significant changes. These have been necessitated by new information, as well as the need to improve clarity. The move from printed versions to the online environment, through the ExplorEnz website, has allowed the process of adding newly reported enzymes to be automated and the information content to be enriched.

Characterization of the in vitro binding and inhibition kinetics of primary amine oxidase/vascular adhesion protein-1 by glucosamine.

Background: Primary-amine oxidase (PrAO) catalyzes the oxidative deamination of endogenous and exogenous primary amines and also functions, in some tissues, as an inflammation-inducible endothelial factor, known as vascular adhesion protein-1. VAP-1 mediates the slow rolling and adhesion of lymphocytes to endothelial cells in a number of inflammatory conditions, including inflammation of the synovium.

Methods: Glucosamine binding to the enzyme was assessed spectrofluorometrically and the kinetics of inhibition of PrAO were determined spectrophotometrically through the use of direct or coupled assays, in the presence of different substrates.

Kinetic behavior and reversible inhibition of monoamine oxidases--enzymes that many want dead.

Monoamine oxidase (MAO) inhibitors have proven to be valuable tools in pharmacology and therapeutics. This account concerns the behavior of the different types of reversible inhibitor and how an understanding of the kinetic mechanisms of MAO may help in their design.

Highly reactive oxygen species: detection, formation, and possible functions.

The so-called reactive oxygen species (ROS) are defined as oxygen-containing species that are more reactive than O(2) itself, which include hydrogen peroxide and superoxide. Although these are quite stable, they may be converted in the presence of transition metal ions, such as Fe(II), to the highly reactive oxygen species (hROS). hROS may exist as free hydroxyl radicals (HO·), as bound ("crypto") radicals or as Fe(IV)-oxo (ferryl) species and the somewhat less reactive, non-radical species, singlet oxygen.

Recommendations for terminology and databases for biochemical thermodynamics.

Chemical equations are normally written in terms of specific ionic and elemental species and balance atoms of elements and electric charge. However, in a biochemical context it is usually better to write them with ionic reactants expressed as totals of species in equilibrium with each other. This implies that atoms of elements assumed to be at fixed concentrations, such as hydrogen at a specified pH, should not be balanced in a biochemical equation used for thermodynamic analysis.