Abstract
The subunit structure and complete amino acid sequence of the lectin extracted from Lens culinaris (LcL) seeds was determined. In previous studies, the primary structure of the alpha-chain (Mr = 5,710) was shown to be homologous to the alpha-chain of the lectin from Pisum sativum, the Vicia cracca glucose-specific lectin, and a region in the middle of the concanavalin A sequence (residues 70-121). The complete amino acid sequence of the beta-chain (Mr = 17,572) has been determined from 11 tryptic peptides, 4 peptides derived by chemical cleavage of the beta-chain at its three tryptophan residues, 11 peptides obtained after digestion with Staphylococcus aureus protease, and 5 tryptic peptides from the succinylated polypeptide chain. The extensive homologies by alignment of the alpha- and beta-chains of the L. culinaris lectin with portions of concanavalin A situated between 1 to 45 and 70 to 237, suggest that the L. culinaris and Canavalia ensiformis lectins have evolved from each other. A comparison was made between the secondary structure of the C. ensiformis lectin and the probable secondary structure of the L. culinaris lectin as predicted by two different methods. The results indicate that the folding of these two polypeptides has been particularly well conserved during evolution. It is suggested that the L. culinaris lectin is synthesized as a single polypeptide chain and cleaved subsequently into two or possibly three fragments, two of which would be alpha and beta and the third a fragment homologous to portion 46 to 69 in concanavalin A. As in favin, the amino acids postulated to be involved in the formation of the hydrophobic cavity and the sugar and metal binding sites are highly conserved in L. culinaris lectin.
Highlights
The subunit structure and complete amino acid se- lectins, in terms of chemical structure andcarbohydrate-bindquence of the lectin extracted from Lens culinaris (LcL) ing properties, are derived from plants [3]
The primary lectins display extensive variations in chemical structure of the a-chain (Mr = 5,710) was shown to be and physical properties,astructure-function relationship, homologous to the a-chain of the lectin from Pisum based on NHz-terminal sequences, has beendemonstrated sativum, the Vicia cracca glucose-specific lectin, and a region in the middle of the concanavalin A sequence among selected lectins from leguminous plant seeds [5]
An attempt canavalin A situated between 1 to 45 and 70 to 237, was made to predict the secondary structure of the lentil lectin suggest that theL. culinaris and Canavalia ensiformis using two different calculation methods [8, 9]
Summary
Determination of the Primary Structureof the LentilLectin-The lentil lectin (LcL) was isolated following the method of Hayman and ‘The abbreviations used are: Con A, ccncanavalin A; LcL, L. culinaris lectin; PTH, phenylthiohydantoin; dansyl-, 5-dimethylaminonaphthalene-1-sulfonyl-. Full size photocopies are included in the microfilm edition of the Journal thatis available from Waverly Press. The complete amino acid sequence of the LcL 8-chain was determined from peptides obtained following tryptic digestion (before and after succinylation), digestion with S. aureus protease, and chemical b cleavage at thetr.yptophan residues. P,, respectively, of the 20 amino acids that havebeen quantitatively determined froma statistical analysisof globular proteinswith known conformation. ( c ) Alarge of agreement with the preceding method In this method, therole of each type of amino acid residue in settling its own conformation and that of its neighbors is correlated withphysicochemical properties of the residue. Number of “SP”-peptides was obtained by digestion with S. aureus protease, an enzymwheich cleaves peptide bonds after glutamic acid andaspartic acidresidues. (d)Tryptic digestion of the native P-chainyielded a number of ‘7“’-peptides which were separated by gel filtra-. Tion. (e) The remaining partsof the ,&chain sequence and the missing overlaps were obtained by sequence determinationof “W” fragments, resulting from CNBr cleavage after tryptophan residues
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