Abstract
Abstract Soybean agglutinin purified by affinity chromatography on Sepharose-e-aminocaproyl-β-d-galactopyranosylamine was shown to be homogeneous on disc gel electrophoresis, ultracentrifugation, and gel filtration. Isoelectric focusing in a sucrose gradient gave a pI of 5.81. The extinction coefficient (A1%280) was found to be 12.8 cm-1. Measurement of the partial specific volume gave a value of 0.745 ml per g. The intrinsic sedimentation coefficient (s020,w) was estimated as 6.0 ± 0.12 S, between pH 2.2 and 10.8, and as 2.15 ± 0.15 S in 0.1% sodium dodecyl sulfate, 8 m urea, and 6 m guanidine hydrochloride. The molecular weight of the agglutinin, determined by sedimentation equilibrium and by gel filtration, was found to be 122,000 ± 1,300 and 120,000 ± 10,000, respectively. Disc gel electrophoresis and gel filtration, both in the presence of sodium dodecyl sulfate, and sedimentation equilibrium in 6 m guanidine hydrochloride gave a subunit molecular weight of 30,000 ± 1,500 and 30,300 ± 400, respectively. Four alanine residues per 120,000 g were found by amino-terminal analysis. It is concluded that the agglutinin is a tetramer composed of identical subunits. Two binding sites for N-acetyl-d-galactosamine were found per 120,000 daltons by equilibrium dialysis and gel filtration, with an association constant K = 3.0 x 104 liter mole-1.
Highlights
Soybean agglutinin purified by affinity chromatography on Sepharose-~-aminocaproyl-~-D-galactopyranosylamine was shown to be homogeneouson disc gel electrophoresis, ultracentrifugation, and gel filtration
The molecular weight of the were used for the dissociation of SBA
To protein ratio, 5:l w/w) and dialyzed against 0.1% sodium The partial specific volume was calculated on the basisof dodecyl sulfate
Summary
Soybean agglutinin purified by affinity chromatography on Sepharose-~-aminocaproyl-~-D-galactopyranosylamine was shown to be homogeneouson disc gel electrophoresis, ultracentrifugation, and gel filtration. To protein ratio, 5:l w/w) and dialyzed against 0.1% sodium The partial specific volume was calculated on the basisof dodecyl sulfate.
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