Abstract

8-Azidoadenine was employed as a photoaffinity probe of the adenine binding site of the seed lectin from lima beans and from Phaseolus vulgaris erythroagglutinin. This compound was shown to (a) bind competitively to the adenine binding site of these lectins and (b) exhibit enhanced binding in the presence of 1,8-anilinonaphthalenesulfonic acid in the same manner as adenine. The presence or absence of 1,8-anilinonaphthalenesulfonic acid during labeling caused no change in the peptide maps of either lectin when digested with trypsin. The peptide maps of each lectin showed one major peak of radioactivity. Sequencing of the corresponding tryptic peptide from lima bean lectin indicated the primary structure to be Val-Leu-Ile-Thr-Tyr-Asp-Ser-Ser-Thr-Lys. The sequence of the labeled peptide isolated from P. vulgaris erythroagglutinin was Thr-Thr-Thr-Trp-Asp-Phe-Val-Gly-Glu-Asn-Glu-Val-Leu-Ile-Thr-Tyr, which corresponded to residues 173-190 of the cDNA-derived sequence (Hoffman, L. M., and Donaldson, D. D. (1985) EMBO J. 4, 883-889). Residues 186-190 (italicized) are identical to the first five amino acids in the lima bean lectin peptide. The peptides are located at the COOH-terminal half of the lectin and show extensive homology with other legume lectins.

Highlights

  • Photoaffinity Labeling of the AdenineBinding Site of the Lectins from LimaBean, Phaseolus lunatus, and the Kidney Bean, Phaseolus vulgaris”

  • Scatchard analysis of equilibrium dialysis results indicated NaAde to be an effective competitor for the adenine binding site of LBL and PHA-E (Fig. 1).This approach allowed the calculation of a dissociation constant for the binding of N3Ade to the lectins (Roberts and Goldstein, 1983)

  • We have attempted to determine the primary structure of the adenine binding site region in two legume lectins

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Summary

Introduction

Photoaffinity Labeling of the AdenineBinding Site of the Lectins from LimaBean, Phaseolus lunatus, and the Kidney Bean, Phaseolus vulgaris”. Probe ofthe adenine binding site of the seed lectin from Recently, it has been observed that many, not all lima beans and from Phaseolus vulgaris erythroagglu- legume lectins, contain a specific high affinity binding tinin. This compound was shown to ( a )bind competi- site for adenine and several of its derivatives such as cytokitively to the adenine binding site of these lectins and ( b )exhibit enhanced binding in the presence of 1,8-. The presence or absence of 1,8-anilinonaphthaleneeulfonic acid during labeling caused no change in the peptide maps of either lectin when digested with trypsin. Tyr-Asp-Ser-Ser-Thr-Lys.The sequence of the labeled reduce binding, alterations at positions 2 and 8 are accompeptide isolated from P. vulgaris erythroagglutinin modated by the protein (Roberts et al, 1986)

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