Isolation and Characterization of a Virus Inhibitor from Spinach (Spinacia oleracea L.)

A virus inhibiting protein (VI) was isolated from spinach (Spinacia oleracea L.). The VI inhibited infections of test plants with plus- and minus-strand RNA viruses. Inoculation of both local lesion and systemic hosts with TMV in the presence of varying amounts of the VI resulted in typical dose response curves for the number of local lesions or the amount of virus respectively. The lowest concentration of VI leading to a significant reduction in the number of local lesions was 0.06 μg/ml. The VI was found to inhibit local lesion formation only when applied within 2–3 h p.i. but still reduced the number of local lesions when applied up to 9 h prior to virus inoculation. The antiviral activity could be attributed to a protein of molecular weight 29,000 dalton with an isoelectric point of 10.3. Its activity was destroyed by heating for 30 min to 70°C. These characteristics resemble those of other virus inhibiting proteins described for members of the order Caryophyllales such as the Phytolacca inhibitor against which a serological relationship was obtained.     

Cholesteryl hemisuccinate alters membrane fluidity, angiotensin receptors, and responses in adrenal glomerulosa cells

We examined the effects of cholesteryl hemisuccinate on membrane fluidity and angiotensin II (AII) actions in bovine adrenal glomerulosa cells. Incubating cells with cholesteryl hemisuccinate decreased membrane fluidity and markedly inhibited AII binding. The effect on binding was characterized by a decrease in AII receptor number. The effects of AII on phosphatidyl inositol turnover and calcium fluxes, proposed intermediaries of AII actions on aldosterone secretion, were less impaired than AII binding by cholesteryl hemisccinate. AII stimulation of aldosterone secretion was preserved despite the decrease in AII binding after cholesteryl hemisuccinate treatment. These results indicate that AII binding can be dissociated from its effects on aldosteronogenesis by a reagent that alters membrane fluidity.     

Multiple Forms of Choline-O-Acetyltransferase in Mouse and Rat Brain: Solubilization and Characterization

Three forms of acetyl coenzyme A: choline-O-acetyltransferase (EC 2.3.1.6, ChAT) have been isolated from mouse and rat forebrain synaptosomes with a 100 mM sodium phosphate (NaP) buffer of pH 7.4, a high-salt solution (500 mM NaCl), and a 2% Triton DN-65 solution, respectively. The Triton-solubilized form of ChAT differed from the other two forms in its capacity to acetylate homocholine, its pH profile, and its sensitivity to denaturation. NaCl-solubilized ChAT could be distinguished from the other two forms with respect to pH profile, sensitivity to inhibition by 4-(1-naphthylvinyl) pyridine (in the presence of Triton), and apparent Km value for choline acetylation. The caudate and putamen of rat brain contained the highest amount of ChAT activity, based on tissue wet weight, and the cerebellum contained the least of the brain regions examined; only the cerebellum had more membrane-bound than soluble ChAT. Septal lesion reduced ChAT activity in the NaP- and Triton-solubilized fractions prepared from hippocampus by 68% and 64%, respectively, whereas it reduced the activity of the NaCl-solubilized fraction by only 21%. These results suggest that three different forms of ChAT may exist in both mouse and rat brain.     

Correlative changes in sucrose uptake, ATPase activity and membrane fluidity in carnation petals during senescence

Apparent sucrose uptake. ATPase activity and membrane fluidity changes were studied during the development and senescence of carnation flowers ( Dianthus caryophyllus L., cv. Cerise Royallette). Typical changes associated with senescence of a cut flower, such as respiration, ethylene production and fresh weight, were measured. Concomitant with a rise in respiration and ethylene production and a decline in fresh weight, a sharp decrease in apparent sucrose uptake was observed. Sucrose uptake was pH dependent (pH optimum, 5.5) and influenced by membrane integrity. Apparently, the activity of ATPase is related to sucrose uptake, because similar changes occurred during flower development. In addition, the activity of ATPase was well correlated with membrane fluidity.
It is suggested that sucrose uptake is controlled by ATPase activity, which in turn is modulated by membrane lipid fluidity. The decline in membrane fluidity associated with senescence leads to a corresponding reduction in ATPase activity and sucrose uptake. Further evidence supporting this view comes from experiments in which senescence was enhanced by 1-aminocyclopropane-l-carboxylic acid. It shortened the time to decline in fresh weight, rise in respiration and ethylene production. In parallel, reduction in membrane fluidity, ATPase activity and sucrose uptake were observed.     

Sequence interrelationships of the subunits of vicilin from pea seeds

Serological studies and comparison of N-terminal amino acid sequences with the amino acid sequence deduced from a cDNA clone have been used to establish the sequence relationships between the subunits of the pea seed storage protein, vicilin. Subunits smaller than M_r～50 000 (i.e., M_r 34 000, 30 000, 25 000, 18 000, 14 000, 13 000 and 12 000) show extensive homology with molecules within M_r～50 000 group. Both the sequencing and serological data confirm earlier evidence from studies on vicilin synthesisin vivo andin vitro which indicated that the vicilin subunits smaller than M_r～50 000 arose by endoproteolytic cleavage of parent molecules within the M_r～50 000 group. Cleavage in different M_r 50 000 parent molecules containing either one or both of two susceptible processing sites accounts for the formation of all the vicilin subunits smaller than M_r～50 000, with the possible exception of the M_r34 000 polypeptide. The position of these sites in the putative parents were defined by reference to a complete amino acid sequence deduced from the sequence of DNA complementary to mRNA for one member of the M_r～50 000 group.     

Separation and properties of human brain hexosaminidase C

Hexosaminidase C was separated from human brain supernatant by immunoadsorption of the A and B forms on to a column of immobilized antibody followed by preparative starch-block electrophoresis. There were some differences in the properties of hexosaminidase C preparations after each of these stages, shown by comparison of their heat-inactivation characteristics and filtration through Bio-Gel P-200. The C form prepared by both separation steps had properties which differed markedly from those of the A and B isoenzymes; its molecular weight was much larger, greater than 200000, it had optimum activity between pH6 and 7 and could not be successfully eluted from DEAE-cellulose, even with high salt concentrations, or from Sephadex G-200. These results seem to support the proposal that the C form is under a separate genetic control from the others.