TY - JOUR
T1 - Mass spectrometric characterization of isoform variants of peanut (Arachis hypogaea) stem lectin (SL-I)
AU - Agrawal, Praveen
AU - Kumar, Saravanan
AU - Das, Hasi Rani
N1 - Funding Information:
Authors wish to thank Prof. R. H. Das, School of Biotechnology, Guru Gobind Singh Indraprastha University, Delhi for going through the manuscript critically. Authors gratefully acknowledge Somdutta Sen and Reena Arora, The Centre for Genomic Application (TCGA), Delhi, for help in interpretation of mass spectrometric data. Praveen Agrawal is thankful to the Council of Scientific and Industrial Research (CSIR), New Delhi, India for awarding the Senior Research Fellowship.
PY - 2010/6/16
Y1 - 2010/6/16
N2 - 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.
AB - 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.
KW - Isoform
KW - Lectin
KW - MS/MS based peptide sequencing
KW - Mass spectrometry
KW - Microheterogeneity
KW - Peptide mass fingerprinting
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U2 - 10.1016/j.jprot.2010.03.006
DO - 10.1016/j.jprot.2010.03.006
M3 - Article
C2 - 20348039
AN - SCOPUS:77953098857
SN - 1874-3919
VL - 73
SP - 1573
EP - 1586
JO - Journal of Proteomics
JF - Journal of Proteomics
IS - 8
ER -