Faan Wen Bangerter

The rapid intermembraneous transfer of retinoids

Abstract The intermembraneous rates of retinoid (all- trans -retinol(al), 11- cis -retinol and all- trans -retinol palmitate) transfer from vesicle to vesicle and vesicle to erythrocyte were studied. The rates of transfer of the retinols(al) were exceedingly rapid. The rates of transfer of the retinols(al) from egg phophatidyl choline based SUVs to bovine erythrocytes had a half-time of approximately 1–2 min. The vesicle to vesicle transfer rate was too rapid to measure by conventional techniques. By contrast, all- trans -retinol palmitate did not undergo transfer at an appreciable rate.

The synthesis and properties of a functional fluorescent cholesterol analog

Abstract The synthesis of a new fluorescent cholesterol analog is described. The analog contains a cholesterol nucleus attached via a hydrophilic spacer to N-4-nitrobenzo-2-oxa-1,3-diazole. Since the cholesterol moiety is not perturbed this molecule probably interacts with lipid bilayers in much the same way as cholesterol itself does. The compound can be readily incorporated into small unilamellar vesicles by sonicating a mixture of it with egg yolk phosphatidylcholine in a buffer. Furthermore, the analog can be incorporated into preformed membranes either by exchange from vesicles containing the analog or by uptake from sonicated micelles of the analog. Thus this analog shows potential as a useful tool for studying the interactions of cholesterol with cell membranes.

The Inactivation of γ-Aminobutyric Acid Transaminase in Dissociated Neuronal Cultures from Spinal Cord

Abstract: It had previously been shown that dissociated cell cultures from chick embryo spinal cord have a high affinity uptake system for the neurotransmitter γ‐aminobutyric acid (GABA) and make functional inhibitory synaptic contacts as determined by electrophysiology (Farb et al., 1979). It is shown here that these cultures can synthesize GABA from added glutamate in a glutamate decarboxylase‐dependent reaction. Furthermore, these cultures have a functional GABA transaminase that degrades the neurotransmitter. This enzyme can be specifically and irreversibly blocked with gabaculine. A 15 min incubation with 10−6 M‐gabaculine completely inactivates the enzyme. The inactivation of the enzyme leads to an increase in GABA levels. Long‐term incubation (16 days) of gabaculine in the medium does not appear to alter high affinity GABA transport, suggesting that the drug is not toxic to cells capable of accumulating GABA.

On the labelling of oxidized cell surface membranes by acyl hydrazides

Abstract The covalent modification of bovine erythrocyte membranes by first oxidizing cell surface sugars followed by the covalent coupling of the oxidized components with hydrazides was investigated. The aldehydic groups were introduced either chemically, by mild periodate oxidation, or enzymically with galactose oxidase. The periodate (0.1 mM) and galactose oxidase (50 units) oxidations were complete at 5 and 240 min, respectively, in phosphate-buffered saline (pH 7.4) at 30°C using 2 · 109 cells · ml−1. The rates of hydrazone formation between the oxidatively generated aldehyde groups and α-[3H]acethydrazide were also measured (pH 7.4) at 30°C. The reactions with the periodate and galactosde oxidase-treated cells went to completion in 1.5 and 6 h, respectively. Aniline had a marked catalytic effect on these rates. With 5 mM aniline under the same conditions, the periodate-treated cells reacted to completion with acethydrazide in 30 min and the galactose oxidase-treated cells in 40 min. As expected, although the rates of α-[3H]acethydrazide incorporation were increased with aniline, the extents of the reaction were not. Finally, the stability of the hydrazone linkages were measured. The hydrazone bonds formed when the galactose oxidase-treated cells were reacted with acethydrazide were completely stable at 30°C for 24 h whereas those formed from the periodate-treated cells were not. In the latter case roughly 50% of the membrane-bound hydrazone linkages were lost after 6 h at 30°C. The remaining 50%, however, were stable to cleavage. Thus, these sites are clearly heterogeneous in nature.

Intermembranous transfer of retinoids

Publisher Summary This chapter discusses the procedures of intermembranous transfer of retinoids. It is generally assumed that intermembranous hydrophobic ligand transfer proteins are reserved for compounds that undergo very slow transfer in the absence of a catalyst. It has been shown that nonesterified retinoids undergo rapid intermembranous transfer in the absence of catalytic transfer or exchange proteins. The rates of transfer are at least an order of magnitude faster than that for cholesterol. These results suggest that carrier protein assistance is not essential for catalyzing bulk transfer of the retinols(als) in the retina and retinal pigment epithelium. The intermembrane transfer techniques discussed in this chapter are of qualitative use for rapid transfer processes such as those found in the case of retinol and retinal. This is because the standard separation techniques are useful only when the half-times for transfer occur on a time scale of minutes, whereas the actual transfer rates for the retinol(al)s occur within seconds.