Distribution of Iron-Containing Superoxide Dismutase in Vascular Plants

Abstract

Superoxide dismutases (EC 1.15.1.1) in vascular plants representing different evolutionary levels were characterized using polyacrylamide gel electrophoresis. The three forms of the enzyme were distinguished from each other based on the following criteria: a) the Cu-Zn enzyme is sensitive to cyanide wherease the Fe and Mn enzymes are not; and b) the Cu-Zn and Fe enzymes are inhibited by H202 whereas the Mn enzyme is H202-resistant. Of the 43 plant families investigated, the Fe-containing superoxide dismutase was found in three families: Gingkoaceae, Nymphaceae, and Cruciferae.yotic algae lack the Cu-Zn SOD but contain either the Mn and/ or Fe enzymes (3,20). Land plants contain large amounts of the Cu-Zn enzyme plus an additional cyanide-resistant SOD which was assumed to be the Mn protein (3,11,17). The Fe enzyme was previously thought to be restricted to procaryotic organisms but has recently been purified from Euglena gracilis (14,18) MATERIALS AND METHODS Mature leaves from native plants and cultivars were collected locally. Etiolated seedlings were grown from seed in a darkened chamber and harvested 7 to 10 days after sowing. Leaves or etiolated seedlings were ground in a Waring Blendor in ice-cold media consisting of 50 mm K-phosphate (pH 7.0), and 0.1% Triton X-100. The homogenate was squeezed through four layers of cheesecloth and centrifuged at 12,000g for 10 min. Leaf extracts were dialyzed overnight against 10 mm K-phosphate (pH 7.0), to remove small mol wt compounds which interfered with the activity stain for SOD on polyacrylamide gels.Electrophoresis was performed on 7.5% acrylamide gels (9). The photochemical method of Beauchamp and Fridovich (6) was used to localize SOD activity on gels. The three types of SOD were distinguished by their different sensitivity to inhibitors (5,8). Inhibitors (1 mim KCN or 2 mi H202) were added to the staining solution prior to addition to the gels. Gels were incubated in the staining solution for 45 min before exposure to light. Densitometric scans of gels were made with a Perkin-Elmer model 552-0800 integrating gel scanner. Spectrophotometric measurements of SOD activity were made by the method of McCord and Fridovich (21). Assays for SOD were routinely done in the presence of 10 ylM KCN to inhibit Cyt oxidase. This low cyanide concentration was insufficient to inhibit the Cu-Zn enzyme. RESULTSThe Cu-Zn enzyme could easily be distinguished from the other two isozymes by its sensitivity to cyanide. Asada et al.

Superoxide Dismutase Enhances Tolerance of Freezing Stress in Transgenic Alfalfa (Medicago sativa L.)

“…Although the relative activity of SOD varied in leaf age and growth conditions among the plants, the transgenics consistently had a 30 to 100% increase in total SOD activity. The inclusion of 100 p~ cyanide-in the assay mixture inhibits Cu/Zn-SOD activity (Bridges and Salin, 1981) and, therefore, estimates Mn-SOD activity. In the presence of cyanide-three of the transgenic plants had approximately 45 to 95% increase in Mn-SOD activity relative to RA3 (Table I), but the stimulation of total SOD in the transgenic plants and the amount of Mn-SOD activity did not mathematically correspond.…”

Synthesis, Characterization and Biological Evaluation of Novel Thiadiazoline Sulfonamides and Metal Complexes

“…The new thiadiazoline sulfonamide derivatives and metal complexes with N-phenyl 2-pyridine carbohydrazonoyl halide were assayed for SOD enzyme-like activity (Bridges and Salin, 1981). The SOD-like activity of the thiadiazoline sulfonamide derivatives and metal complexes with N-phenyl 2-pyridine carbohydrazonoyl halide was assayed using phenazine methosulfate to generate superoxide anion radicals at pH = 8.3 (phosphate buffer).…”

Synthesis and Biological Activity of Drug Delivery System Based on Chitosan Nanocapsules

“…The chitosan, naproxen and naproxen/chitosan nanocapsule were assayed for super oxide dismutase enzyme like activity [34]. SOD like activity of chitosan, naproxen and naproxen/chitosan nanocapsule was assayed by using phenazenmethosulphate (PMS) to generate a superoxide anion radicals at pH = 8.3 (phosphate buffer).…”