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Organelles of eukaryotic cells are structures made up of membranes, which carry out a majority of functions necessary for the surviving of the cell itself. Organelles also differentiate the prokaryotic and eukaryotic cells, and are arranged to form different compartments guaranteeing the activities for which eukaryotic cells are programmed. Cell membranes, containing organelles, are isolated from cancer cells and erythrocytes and used to form biocompatible and long circulating ghost nanoparticles delivering payloads or catalyzing enzymatic reactions as nanoreactors. In this attempt, red blood cell membranes were isolated from erythrocytes, and engineered to form nanoerythrosomes (NERs) of 150 nm. The horseradish peroxidase, used as an enzyme model, was loaded inside the aqueous compartment of NERs, and its catalytic reaction with Resorufm was monitored. The resulting nanoreactor protected the enzyme from proteolytic degradation, and potentiated the enzymatic reaction in situ as demonstrated by maximal velocity (V-max) and Michaelis constant (K-m), thus suggesting the high catalytic activity of nanoreactors compared to the pure enzymes.

Lipases from Rhizomucor miehei (RmL) and Candida antartica (CaL-B) entrapped in surfactant free microemulsion-like ternary systems consisting of n-hexane, short chain alcohols (1-propanol or 2-methyl-2-propanol) and water, keep their catalytic activity at 30 degrees C, catalyzing efficiently the esterification of fatty acids or natural phenolic acids including cinnamic acid derivatives. High conversions (up to 95%) were obtained with both lipases entrapped in various ternary systems. Lipases catalytic efficiency and stability was influenced by the composition of the ternary systems used. Hydroxypropylmethyl cellulose (HPMC) microemulsions-based organogels (MBGs) formulated with various surfactant free microemulsion-like systems were used as a matrix for enzyme immobilization. These lipase-containing MBGs prove to be a novel solid-phase catalyst for use, under mild conditions, in non-polar solvents as well as in solvent free systems. CaL-B immobilized in surfactant free MBGs keep its catalytic activity for a storage period of 8 months, while the corresponding residual activity of immobilized RmL was decreased by 50%. The stability of enzymes in such surfactant free MBGs is higher than that observed in MBGs formulated with surfactant based w/o microemulsions. Lipases-containing surfactant free MBGs were used in seven consecutive batch syntheses of fatty esters with only 10-15% loss of activity. (c) 2005 Elsevier Inc. All rights reserved. Enzyme and Microbial Technology

Modeling methods allow the identification and analysis of determinants of reactivity and specificity in enzymes. The reaction between glutathione and 1-chloro-2,4-dinitrobenzene (CDNB) is widely used as a standard activity assay for glutathione S-transferases (GSTs). It is important to understand the causes of differences between catalytic GST isoenzymes and the effects of mutations and genetic polymorphisms. Quantum mechanical/molecular mechanical (QM/MM) molecular dynamics simulations have been performed here to investigate the addition of the glutathione anion to CDNB in the wild-type M1-1 GST isoenzyme from rat and in three single point mutant (Tyr6Phe, Tyr115Phe, and Met108Ala) M1-1 GST enzymes. We have developed a specifically parameterized QM/MM method (AM1-SRP/CHARMM22) to model this reaction by fitting to experimental heats of formation and ionization potentials. Free energy profiles were obtained from molecular dynamics simulations of the reaction using umbrella sampling and weighted histogram analysis techniques. The reaction in solution has also been simulated and is compared to the enzymatic reaction. The free energies are in excellent agreement with experimental results. Overall the results of the present study show that QM/MM reaction pathway analysis provides detailed insight into the chemistry of GST and can be used to obtain mechanistic insight into the effects of specific mutations on this catalytic process.