Structural Studies on Antigen E of Ragweed Pollen

Abstract

The analysis of antigen E of ragweed pollen by polyacrylamide gel electrophoresis in sodium dodecyl sulfate (SDS) resulted in the demonstration of two subunits of molecular weights of about 20 000 and 14 500. These results confirm earlier studies on antigen E by gel chromatography that demonstrated a two-polypeptide subunit composition. Treatment of antigen E with 1% SDS (in the absence of reducing agent) resulted in the dissociation of the two polypeptide chains, which indicates that the bonds bridging the subunits are non-covalent in nature.

Studies of energy‐linked reactions: Dihydrolipoate‐ and oleate‐ dependent ATP synthesis in yeast promitochondria

“…Previous experiments indicate that the respiratory chain is not necessary for dihydrolipoate-dependent ATP synthesis [3,4] as the reaction occursin the presence of rotenone and antimycin A and the present experiments with promitochondria appear to confirm that the cytochromes and ubiquinone are not involved. However, the redox reactions involved in dihydrolipoate-dependent ATP synthesis have not been characterised nor has the redox couple which is the electron acceptor in this reaction.…”

“…where a specific requirement for unsaturated fatty acids has been demonstrated [3]. ATP synthesis in promitochondria from UFA-depleted yeast cells can be restored by addition of oleic acid and oleoyl CoA (table 3) as is found in the case of detergent-dispersed ATP synthase preparations.…”

“…A possible role for ATP-generating reactions in promitochondria and in yeast 'petite' mitochondria has been discussed by Kovac [lo] on the basis of sensitivity to inhibitors and studies of cytochrome de& cient mutants. Detailed examination of dihydrolipoatedependent, oleoyl-S-lipoate-dependent and oleoyl phosphate-dependent ATP synthesis [3,4] in mitochondrial 'petite' mutants, nuclear 'petite' mutants and mitochondrial mitmutants with specific deletions in cytochrome content and ATPase subunits should be of value in determining the components involved as well as establishing their biogenetic origin.…”

“…DLa-lipoic acid was dissolved in methanol. Oleoyl-S-lipoate was prepared from dihydrolipoate and oleoyl chloride as described previously [3] and dissolved in dimethylformamide.…”

“…Total promitochondrial fatty acids were determined by gas chromatography of the methyl esters, methylation being carried out in 0.5 M HCl-methanol at 60°C for 2 h. The methyl esters were analysed on a Perkin-Elmer F-l 1 FEBSLETTERS February 1977 instrument with a polyethylene glycol succinate column operating at 180°C. ATP synthesis was assayed in a glucose-hexokinase trap system by the disappearance of inorganic phosphate [3]. The reaction medium contained 1 mg promitochondrial protein, 250 mM sucrose, 22 mM glucose, 2 mM MgCl?, 0.2 mM EDTA, 2 mM ADP, 20 mM Tris-Cl, pH 7.3 and 25 units of hexokinase.…”