Electron-transferring enzymes in the plasma membrane of the Ehrlich ascites tumor cell

Abstract

The plasma membrane of the Ehrlich ascites tumor cell contains an NADH dehydrogenase. This activity was shown not to be due to contamination by other subcellular membranes. A variety of electron acceptors have been compared as to rate with the following result: ferricyanide greater than cytochrome c greater than cytochrome b5 greater than glyoxylate greater than dichlorophenolindophenol. Oxygen acceptance could not be detected. The optimum assay temperature and pH ranges were 30--40 degrees C and pH 6--8, respectively. With respect to either NADH or ferricyanide, the kinetics yielded linear double-reciprocal plots. Inhibition of the enzyme by sulfhydryl reagents could be blocked by excess NADH. Detergents such as Triton X-100 or cholate resulted in solubilization of the enzymatic activity, but phospholipase A2 did not. The activity differed from that of the mitochondria in that it was not inhibited by rotenone or antimycin A. The possible involvement of NADH oxidation in the energetics of plasma membrane transport is discussed.

Tumor necrosis factor stimulates amino acid transport in plasma membrane vesicles from rat liver.

“…Membrane pellets were resuspended in SMB and to an approximate protein concentration of 8-10 mg/ ml and aliquots were stored at -70'C in cryotubes (InterMed; Nunc, Roskilde, Denmark) until studied. Vesicle purity was evaluated biochemically by assaying the relative enrichment or impoverishment in the specific activity of the marker enzymes 5 '-nucleotidase (21 ), glucose-6-phosphatase (22), and NADPH/cytochrome c reductase (23). Inorganic phosphate was measured according to Fiske and Subbarow (24).…”

Plant polyphenols as electron donors for erythrocyte plasma membrane redox system: validation through in silico approach

“…Later researches established the existence of trans-membranous NADH dehydrogenases in several other cell types [2,3]. Evidence is now clear for the presence of a trans-plasma membrane electron transport or plasma membrane redox system (PMRS) in all organisms including bacteria, yeast, animals and plants [4,5].…”

Sodium-dependentl-serine transport in plasma membrane vesicles isolated from Ehrlich cells by two-phase compartmentation

“…Several different methods are described (Colombini & Johnstone, 1973;Kilberg & Christensen, 1979;Im & Spector, 1980;Medina, 1989) which suffer from slowness, poor recovery, and/or contamination by other subcellular fractions. The good results obtained with the method used in the present work reveal the great capabilities of 2-phase compartmentation to clearly and efficiently separate different subcellular fractions (Flanagan, 1985).…”

“…The enrichment of plasma membranes obtained with gradient centrifugation methods is usually less (Van Amelsvoort, Sips & Van Dam, 1978;Zafra & Gimdnez, 1986). Previously reported procedures to isolate plasma membrane vesicles from Ehrlich ascites tumor cells yield approximately a 20-fold enrichment in the plasma membrane fraction (Colombini & Johnstone, 1973;Kilberg & Christensen, 1979;Im & Spector, 1980). However, the degree of contamination by internal membranes (especially, mitochondrial membranes) in these preparations is greater than that obtained by the twophase system compartmentation used in the present work.…”