Pepsin degradation of Cry1A(b) protein purified from genetically modified maize (Zea mays).
The aim of this work was to study the in vitro digestion of Cry1A(b) protein by pepsin. To perform this work, a protein fraction purified from transgenic maize by immunoadsorption was employed. The undigested fraction showed several bands of molecular weight ranging between 14 and 70 kDa when assayed by SDS-PAGE. These bands were identified as corresponding to Cry1A(b) protein by immunochemical techniques and mass spectrometry. The rate of degradation of the purified fraction by pepsin estimated by ELISA was found to be about 75% within 30 min, and the protein concentration remained constant up to 4 h. In all treated samples, the full-length protein and fragments present in Cry1A(b) fraction were absent and peptides of less than 8.5 kDa were mainly found by SDS-PAGE and mass spectrometry. These peptides did not react with antiserum against Cry1A(b) protein by Western blotting. These results suggest that Cry1A(b) fraction purified from transgenic maize is rapidly and extensively degraded by pepsin, giving peptides of low molecular mass.
de Luis
Ruth
R
Tecnologia de los Alimentos, Facultad de Veterinaria, Universidad de Zaragoza, Miguel Servet, 177, 50013-Zaragoza, Spain.
Lavilla
María
M
Sánchez
Lourdes
L
Calvo
Miguel
M
Pérez
María D
MD
D000821:Animal Feed; D000818:Animals; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015153:Blotting, Western; D004063:Digestion; D004591:Electrophoresis, Polyacrylamide Gel; D004731:Endotoxins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D005753:Gastric Mucosa / Q000378:metabolism; D006460:Hemolysin Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010434:Pepsin A / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D018515:Plant Leaves / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics* / Q000378:metabolism; D011817:Rabbits; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013552:Swine; D003313:Zea mays / Q000235:genetics / Q000378:metabolism*
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D010446:Peptide Fragments; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D010434:Pepsin A