[Implications of the incorporation of transisomers of unsaturated fatty acids into cell membranes]. / Implications de l'incorporation d'isomeres trans d'acides gras insaturés au niveau des membranes cellulaires.

The incorporation of dietary trans fatty acids is shown into intestinal brush border and heart mitochondrial membranes obtained from rats fed with either partially hydrogenated soybean oil or with a synthetic triglyceride containing elaidic or linelaidic acid. The distribution of dietary linelaidate in the major phospholipid classes from inner membranes of mitochondria is determined. Heart mitochondria from normal rats and linelaidic acid fed rats are compared with respect to their swelling rate and their oxidative phosphorylation. The influence of dietary linelaidate on the activity of some brush border-associated enzymes is also studied. On the basis of the authors' observations and those made by others with microorganism cels or models of biological membranes (liposomes and monomolecular films), the possible implications of trans fatty acids incorporation on membrane functions are reviewed.

Further studies of mode of action of lipolytic enzymes.

Pancreatic lipase and phospholipase A2 have been shown by the monomolecular film technique to be progressively inactivated when adsorbed at the interface of their respective substrates. This inactivation is faster for lipase than for phospholipase. It is also enhanced by low film pressures and film transfer. The use of radioactive phospholipase and lipase samples offered the possibility to measure the amount of enzyme adsorbed at a monomolecular film with a reasonable accuracy. This adsorption was found to be relatively slow under the conditions of the assays. The main conclusion drawn from these data is that the enzyme kinetics in presence of a substrate film, and probably also under bulk conditions, is controlled by an adsorption flux responsible for an initial lag period and an inactivation flux tending to decrease the reaction rate. The kinetics are linear only when both fluxes equilibrate.

Effects of colipase on hydrolysis of monomolecular films by lipase.

In a system free of bile salts we measured lipase hydrolysis of 1,3-didecanoylglycerol films in the presence or absence of colipase at different surface pressures. The strong, but not absolutely specific protective effect of colipase, most visible at low surface pressure, can account for the higher enzyme activity in the presence of colipase. This can be understood by taking into account simultaneous penetration and surface inactivation fluxes. Using radioactively labeled lipase, we have shown for the first time in a bile salt-free system that the critical surface pressure above which lipase can no longer penetrate a 1,2-didodecanoylphosphatidylglycerol monlayer is around 23 dynes/cm. Colipase increased this critical surface pressure to 30 dynes/cm indicating that it enables lipase penetration between 23 and 30 dynes/cm. The transfer experiment showed that colipase acts by first penetrating the lipid film and then serving as an anchor for lipase into the film.

Comparative studies of lipase and phospholipase A2 acting on substrate monolayers.

The kinetic aspects of lipolysis by pancreatic lipase and phospholipase A2 from different sources have been compared using monomolecular films of short chain lipids as the substrates. Phosphatidylcholine monolayers, in contrast to phosphatidylethanolamine and phosphatidylglycerol monolayers, were resistant to hydrolysis by pancreatic lipase. The induction time, measured during pre-steady state conditions, increased abruptly for a given value of the surface pressure. This appears to be due to a degree of lipid packing above which the enzyme no longer can penetrate the lipid film. The existence of an optimum in the velocity versus surface pressure profile is the result of at least two counterbalancing factors. As the surface pressure increases, the amount of enzyme present in the interface decreases, whereas the minimal specific activity of the enzyme increases. From this study with monolayers we can conclude that activity of lipolytic enzymes used as tools for probing biological membranes will be greatly influenced by the physiochemical nature of the membrane-water interface. Thus, studies such as this one which can measure the penetrating ability of various lipolytic enzymes can be useful in deriving a better understanding of biological membrane structure.