Lipid Breakdown in Smooth Microsomal Membranes from Bean Cotyledons Alters Membrane Proteins and Induces Proteolysis

The effects of lipid degradation on proteins of smooth microsomalmembranes isolated from young bean cotyledons have been examinedby three techniques, viz. fluorescence energy transfer fromtryptophan to cis-parinaric acid; protein spin-labelling with3-maleimido PROXYL; and SDS-PAGE. Lipid degradation was inducedin isolated membranes by activating phospholipase D and phosphatidicacid phosphatase through the addition of Ca²⁺, by treatmentwith exogenous phospholipase C to simulate the concerted actionsof phospholipase D and phosphatidic acid phosphatase or by treatmentwith exogenous phospholipase A₂ to generate endogenous substratefor lipoxygenase. All of the treatments induced time-dependentchanges in lipid-protein interaction and in protein conformation,and the treatment with phospholipase A₂ also engendered proteolysis.The effects of the Ca²⁺ and phospholipase C treatments on lipid-proteininteraction and protein conformation can presumably be partlyattributed to an accumulation of diacylglycerol in the membrane,whereas the induction of proteolysis by phospholipase A₂ appearsto be due to activated oxygen derived from the lipoxygenasereaction and ensuing lipid peroxidation. Lipid degradation inducedby these treatments simulates that which occurs during naturalsenescence of the cotyledons and hence these observations suggestthat loss of protein function and proteolysis in senescing membranesis facilitated by lipolytic and peroxidative activity withinthe lipid bilayer. Key words: Activated oxygen, lipids, membranes, proteins, senescence     

A Microperfusion Chamber for Studying the Growth of Bacterial Cells

A microperfusion chamber for studying the growth of bacterial cells is described. It can be used with unmodified phase contrast systems. Both temperature and the composition of the growth medium can be changed by the operator during observation of bacteria. The use of the chamber is illustrated with observations on the effect of penicillin on Arthrobacter globiformis .