Molecular dynamics simulations of phospholipid bilayers with cholesterol.

To investigate the microscopic interactions between cholesterol and lipids in biological membranes, we have performed a series of molecular dynamics simulations of large membranes with different levels of cholesterol content. The simulations extend to 10 ns, and were performed with hydrated dipalmitoylphosphatidylcholine (DPPC) bilayers. The bilayers contain 1024 lipids of which 0-40% were cholesterol and the rest DPPC.

Cell and process design for targeting of recombinant protein into the culture medium of Escherichia coli.

This paper is a review of strategies to introduce protein into the liquid medium of Escherichia coli K-12 industrial production cells. The cell design strategies are generally based on one of two general mechanisms. The first strategy involves a two-stage translocation using active transporters in the cytoplasmic membrane followed by passive transport through the outer membrane.

Mechanism of dioxygen cleavage in tetrahydrobiopterin-dependent amino acid hydroxylases.

The reaction mechanism for the formation of the hydroxylating intermediate in aromatic amino acid hydroxylases (i.e., phenylalanine hydroxylase, tyrosine hydroxylase, tryptophan hydroxylase) was investigated by means of hybrid density functional theory.

Limiting factors in Escherichia coli fed-batch production of recombinant proteins.

Fed-batch production of recombinant beta-galactosidase in E. coli was studied with respect to the specific growth rate at induction. The cultivations were designed to induce protein production by IPTG at a glucose feed rate corresponding to high mu = 0.

Interfacial orientation of Thermomyces lanuginosa lipase on phospholipid vesicles investigated by electron spin resonance relaxation spectroscopy.

The binding orientation of the interfacially activated Thermomyces lanuginosa lipase (TLL, EC 3.1.1.

Catalytic mechanism of matrix metalloproteinases: two-layered ONIOM study.

The two-layered ONIOM(B3LYP:MNDO) method has been used to investigate the hydrolytical mechanism of matrix metalloproteinases (MMPs), a large family of zinc-dependent endopeptidases capable of degrading a wide range of macromolecules of the extracellular matrix. Human stromelysin-1 (MMP-3) was chosen as a physiologically important member of the MMP family. As a structural reference, X-ray data on the stromelysin-1 catalytic domain (SCD) complexed to the transition state analogue diphenyl piperidine sulfonamide inhibitor was used.

A density functional study of O-O bond cleavage for a biomimetic non-heme iron complex demonstrating an Fe(v)-intermediate.

Density functional theory using the B3LYP hybrid functional has been employed to investigate the reactivity of Fe(TPA) complexes (TPA = tris(2-pyridylmethyl)amine), which are known to catalyze stereospecific hydrocarbon oxidation when H(2)O(2) is used as oxidant. The reaction pathway leading to O-O bond heterolysis in the active catalytic species Fe(III)(TPA)-OOH has been explored, and it is shown that a high-valent iron-oxo intermediate is formed, where an Fe(V) oxidation state is attained, in agreement with previous suggestions based on experiments. In contrast to the analogous intermediate [(Por.

Density functional models of the mechanism for decarboxylation in orotidine decarboxylase.

The mechanism of orotidine 5-monophosphate decarboxylase (ODCase) has been modeled using hybrid Density Functional Theory (B3LYP functional). The main goal of the present study was to investigate if much larger quantum chemical models of the active site than previously used could shed new light on the mechanism. The models used include the five conserved amino acids expected to be the most important ones for catalysis.

Quantum chemical studies of manganese centers in biology.

During the past five years, hybrid density functional theory has been used to study mechanisms for redox-active enzymes containing complexes with a variety of different transition metals. In this paper, suggested mechanisms of some manganese enzymes are described. For photosystem II, a mechanism is proposed leading to an oxyl radical in the S(3)-state, which is the precursor for the O-O bond formation.

A theoretical study of the mechanism for peptide hydrolysis by thermolysin.

The catalytic mechanism for peptide hydrolysis by thermolysin has been investigated using the B3LYP hybrid density functional method. The starting structure for the calculations was based on the X-ray crystal structure of the enzyme inhibited with the ZF (p)LA phosphonamidate transition-state analogue. Besides the three Zn ligands His142, His146 and Glu166, a few additional residues were also included in the model.

Growth rate-dependent changes in Escherichia coli membrane structure and protein leakage.

The phospholipid and fatty acid content of the Escherichia coli membrane were investigated during continuous cultivation. At low growth rates, there was an increase in cardiolipin produced at the expense of phosphatidylethanolamine. Phosphatidylglycerol had a maximum at a growth rate of 0.

Size as a parameter for solvent effects on Candida antarctica lipase B enantioselectivity.

Changes in solvent type were shown to yield significant improvement of enzyme enantioselectivity. The resolution of 3-methyl-2-butanol catalyzed by Candida antarctica lipase B, CALB, was studied in eight liquid organic solvents and supercritical carbon dioxide, SCCO(2). Studies of the temperature dependence of the enantiomeric ratio allowed determination of the enthalpic (Delta(R-S)Delta H(++)) as well as the entropic (Delta(R-S)Delta S(++)) contribution to the overall enantioselectivity (Delta(R-S)Delta G(++)= -RTlnE).

Stable linker peptides for a cellulose-binding domain-lipase fusion protein expressed in Pichia pastoris.

Fusion proteins composed of a cellulose-binding domain from Neocallimastix patriciarum cellulase A and Candida antarctica lipase B were constructed using different linker peptides. The aim was to create proteolytically stable linkers that were able to join the functional modules without disrupting their function. Six fusion variants containing linkers of 4-44 residues were expressed in Pichia pastoris and analysed.