Biosynthesis, primary structure and molecular cloning of snowdrop (Galanthus nivalis L.) lectin.
Poly(A)-rich RNA isolated from ripening ovaries of snowdrop (Galanthus nivalis L.) yielded a single 17-kDa lectin polypeptide upon translation in a wheat-germ cell-free system. This lectin was purified by affinity chromatography. Translation of the same RNA in Xenopus leavis oocytes revealed a lectin polypeptide which was about 2 kDa smaller than the in vitro synthesized precursor, suggesting that the oocyte system had removed a 2-kDa signal peptide. A second post-translational processing step was likely to be involved since both the in vivo precursor and the Xenopus translation products were about 2 kDa larger than the mature lectin polypeptide. This hypothesis was confirmed by the structural analysis of the amino acid sequence of the mature protein and the cloned mRNA. Edman degradation and carboxypeptidase Y digestion of the mature protein, and structural analysis of the peptides obtained after chemical cleavage and modification, allowed determination of the complete 105 amino acid sequence of the snowdrop lectin polypeptide. Comparison of this sequence with the deduced amino acid sequence of a lectin cDNA clone revealed that besides the mature lectin polypeptide, the lectin mRNA also encoded a 23 amino acid signal-sequence and a C-terminal extension of 29 amino acids, which confirms the results from in vitro translation experiments.
Van Damme
E J
EJ
Laboratory for Phytopathology and Plant Protection, Catholic University of Leuven, Belgium.
Kaku
H
H
Perini
F
F
Goldstein
I J
IJ
Peeters
B
B
Yagi
F
F
Decock
B
B
Peumans
W J
WJ
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System; D003001:Cloning, Molecular; D027801:Galanthus; D037102:Lectins; D037241:Mannose-Binding Lectins; D008861:Microsomes / Q000378:metabolism; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism; D037121:Plant Lectins; D010944:Plants / Q000737:chemistry* / Q000378:metabolism; D011061:Poly A / Q000235:genetics; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000737:chemistry / Q000302:isolation & purification; D012313:RNA / Q000235:genetics; D012333:RNA, Messenger / Q000302:isolation & purification / Q000378:metabolism; D014162:Transfection; D014982:Xenopus laevis
D037102:Lectins; D037241:Mannose-Binding Lectins; D037121:Plant Lectins; D011498:Protein Precursors; D012333:RNA, Messenger; C063117:snowdrop lectin; D011061:Poly A; D012313:RNA