C-terminal post-translational proteolysis of plant lectins and their recombinant forms expressed in Escherichia coli. Characterization of "ragged ends" by mass spectrometry.
Electrospray mass spectrometry was used to accurately measure the molecular masses of single chain lectins from legume seeds and also of three recombinant lectins, expressed in Escherichia coli. The five single chain lectins, Erythrina corallodendron lectin, soybean and peanut agglutinins, Dolichos biflorus lectin, and Phaseolus vulgaris hemagglutinin E, all showed evidence of C-terminal proteolytic processing, in some cases to "ragged" ends, when their masses were compared to those expected from their cDNA sequences and their known carbohydrate chains. Recombinant forms of the lectins from E. corallodendron, soybean, and peanut also showed C-terminal trimming, but not to the same points as the natural forms. Discrepancies between the protein and cDNA sequences of the E. corallodendron lectin were resolved by combined liquid chromatography-mass spectrometry peptide mapping and protein sequencing experiments, and the presence of a second glycosylation site was demonstrated. Our data show that all of these lectins undergo C-terminal proteolytic processing of a readily attacked peptide segment. This trimming is frequently imprecise, and the resulting heterogeneity may be a major contributor to the appearance of isolectin forms of these proteins.
Institute for Biological Sciences, National Research Council of Canada, Ottawa.
N M
NM
Young
D C
DC
Watson
M
M
Yaguchi
R
R
Adar
R
R
Arango
E
E
Rodriguez-Arango
N
N
Sharon
P K
PK
Blay
P
P
Thibault
D000595:Amino Acid Sequence; D002240:Carbohydrate Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D006868:Hydrolysis; D037102:Lectins / Q000235:genetics / Q000378:metabolism*; D013058:Mass Spectrometry / Q000379:methods; D008969:Molecular Sequence Data; D037121:Plant Lectins; D010944:Plants / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D037102:Lectins; D037121:Plant Lectins; D011994:Recombinant Proteins