Mass spectrometric characterization of isoform variants of peanut (Arachis hypogaea) stem lectin (SL-I)

Matrix assisted laser desorption/ionization–time-of-flight (MALDI–TOF) mass spectrometric (MS) analysis of purified Arachis hypogaea stem lectin (SL-I) and its tryptic digests suggested it to be an isoformic glucose/mannose binding lectin. Two-dimensional gel electrophoresis of SL-I indicated six isoforms (A1–A6), which were confirmed by Western blotting and MALDI–TOF MS analysis. Comparative analysis of peptide mass spectra of the isoforms matched with A. hypogaea lectins with three different accession numbers (Q43376_ARAHY, Q43377_ARAHY, Q70DJ5_ARAHY). Tandem mass spectrometric (MS/MS) analysis of tryptic peptides revealed these to be isoformic variants with altered amino acid sequences. Among the peptides, the peptide T12 showed major variation. The ¹⁹⁹Val–Ser–Tyr–Asn²⁰² sequence in peptide T12 of A1 and A2 was replaced by ¹⁹⁹Leu–Ser–His–Glu²⁰² in A3 and A4 (T12′) while in A5 and A6 this sequence was ¹⁹⁹Val–Ser–Tyr–Val²⁰² (T12″). Peptide T1 showed the presence of ¹⁰Asn in the isoforms A1–A5 while in A6 this amino acid was replaced by ¹⁰Lys (T1′). Overall amino acid sequence as identified by MS/MS showed a high degree of similarity between A1, A2 and among A3, A4, A5. Carbohydrate binding domain and adenine binding site seem to be conserved.