Effect of oxygen on the dehalogenation of 1,2-Dibromo-3-chloropropane by cytochrome P450cam (CYP101).

Cytochromes P450 are known to exhibit diverse catalytic functions against a large number of hydrocarbon substrates. The determinants of specific activity(ies) that operate on specific substrates have not been widely explored. Earlier, we showed that dehalogenation of 1,2-dibromo-3-chloropropane (DBCP) by P450cam (CYP101) monooxygenase exhibits oxygen- and substrate-dependent product distributions and reaction rates (1).

A stopped-flow kinetic study of the assembly of nonviral gene delivery complexes.

Stopped-flow circular dichroism and fluorescence spectroscopy are used to characterize the assembly of complexes consisting of plasmid DNA bound to the cationic lipids dimethyldioctadecylammonium bromide and 1, 2-dioleoyl- 3-trimethylammonium-propane and a series of polyamidoamine dendrimers. The kinetics of complexation determined from the stopped-flow circular dichroism measurements suggests complexation occurs within 50 ms. Further analysis, however, was precluded by the presence of mixing (shear) artifacts.

Structure/function relationships of polyamidoamine/DNA dendrimers as gene delivery vehicles.

PAMAM dendrimers are members of a class of polyamine polymers that demonstrate significant gene delivery ability. In this study, a selection of PAMAM dendrimers, spanning a range of sizes (generations 2, 4, 7, and 9) and transfection efficiencies, are characterized by various biophysical methods to search for structural properties that correlate with transfection. Measurements of colloidal properties (size and zeta potential) as a function of charge ratio reveal that highly transfecting dendrimer/DNA complexes have size/zeta potential values between 4 and 8.

Compositional effects of cationic lipid/DNA delivery systems on transgene expression in cell culture.

Studies of the contribution of various physical properties of cationic lipid/DNA complexes (CLDCs) to their observed transgene expression in vitro were conducted using cationic liposomes composed of the cationic lipids 1,2-dioleoyl-3-trimethylammonium propane (DOTAP) and dimethyldioctadecylammonium bromide (DDAB), with or without equimolar amounts of cholesterol (CHOL) or 1,2-dioleoylphosphatidylethanolamine (DOPE). The relative degree of luciferase expression by CLDCs is dependent on a complex relationship between net charge of the CLDC as well as previously reported properties, such as membrane fluidity and curvature of the cationic bilayer. Assessments were made of the role of these physical properties on CLDC stability in the extracellular medium, the extent of DNA cellular association, and membrane disruption activity.

Biophysical characterization of PEI/DNA complexes.

The main goal of this study was to determine the effects of polyethylenimine (PEI) molecular weight and structure (750 kDa, 25 kDa, 2 kDa branched, and 25 kDa linear PEI) and the nitrogen/phosphate (N/P) molar ratio on the physical properties and transfection efficiencies of PEI/DNA complexes. Fourier transform infrared spectroscopy revealed that DNA remained in the B conformation when complexed to all PEIs. Unique alterations in the circular dichroism spectra of DNA were observed in the presence of each PEI, whereas differential scanning calorimetry measurements showed that all PEIs examined destabilized supercoiled DNA at N/P < 3/1, but not at higher ratios.