Characterization and quantitation of concanavalin A binding by plasma membrane enriched fractions from soybean root
The binding of concanavalin A (Con A) to soybean root membranes in plasma membrane enriched fractions (recovered from the 34/45% interface of simplified discontinuous sucrose density gradients) was studied using a radiochemical assay employing tritated (/sup 3/H)-Con A. The effect of lectin concentration, time, and membrane protein concentration on the specific binding of /sup 3/H-Con A by the membranes was evaluated. Kinetic analyses showed that Con A will react with membranes in that fraction in a characteristic and predictable manner. The parameters for an optimal and standard binding assay were established. Maximal binding occurred with Con A concentrations in the range of 8 to 16% of the total membrane protein with incubation times greater than 40 min at 22 C. Approximately 10/sup 15/ molecules of /sup 3/H-Con A were bound per microgram of membrane protein at saturation. Binding was reversible. Greater than 92% of the total Con A bound at saturation was released by addition of ..cap alpha..-methyl mannoside. A major peak of /sup 3/H-Con A binding was also observed in fractions recovered from the 25/30% interface of a complex discontinuous sucrose density gradient when membranes were isolated in the absence of Mg/sup 2 +/. When high Mg/sup 2 +/ was present in the isolation and gradient media, the peak was shifted to a fraction recovered from the 34/38% sucrose interface. These results suggest that Con A binding sites are also present on membranes of the endoplasmic reticulum. The amount of Con A bound by endoplasmic reticulum membranes was at least twice the amount bound by membranes in plasma membrane enriched fractions when binding was compared on a per unit membrane protein basis. In contrast, mitochondrial inner membranes, which equilibrate at the same density as plasma membranes, had little ability to bind the lectin.
- Research Organization:
- Univ. of California, Davis
- OSTI ID:
- 5632045
- Journal Information:
- Plant Physiol.; (United States), Journal Name: Plant Physiol.; (United States) Vol. 68:5; ISSN PLPHA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
550301* -- Cytology-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AGGLUTININS
ANTIBODIES
BIOCHEMICAL REACTION KINETICS
CELL CONSTITUENTS
CELL MEMBRANES
CHEMICAL ACTIVATION
CHEMICAL ANALYSIS
CONCANAVALIN
DATA
ENDOPLASMIC RETICULUM
EXPERIMENTAL DATA
GLYCINE HISPIDA
HEMAGGLUTININS
INFORMATION
ISOTOPE APPLICATIONS
KINETICS
LABELLED COMPOUNDS
LEGUMINOSAE
MEMBRANES
NUMERICAL DATA
ORGANOIDS
PLANTS
QUANTITATIVE CHEMICAL ANALYSIS
RADIOCHEMICAL ANALYSIS
REACTION KINETICS
ROOTS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
UPTAKE