Lectin isolation and detection of N-glycoproteins bearing sialic acid and L-fucose residues in human colorectal mucosa and in adenocarcinoma biopsies.
A method to improve the reactivity to specific lectins of N-glycoproteins isolated from human colorectal mucosa and from adenocarcinoma biopsies was developed using a combination of techniques. Total protein extracts were subjected to affinity chromatography, using the immobilised lectin Concanavalin A coupled to Sepharose, by fast performance liquid chromatography (FPLC). N-glycoprotein enriched fractions were resolved by SDS-PAGE, transferred to PVDF membranes and incubated with various lectins. Digoxigenin-conjugated SNA I and MAA lectins were used to detect sialic acid residues. Biotin-conjugated UEA I lectin was used to detect L-fucose residues. By this method, lectin-binding N-glycoproteins were found in a broad relative molecular mass (Mr) range (from 47 to 205 kDa). No tissue-specific N-glycoproteins were observed when human colorectal mucosa and adenocarcinoma samples were compared.
De Carlos
Alejandro
A
Facultad de Ciencias, Universidad de Vigo, E-36200 Vigo, Spain. adcarlos@uvigo.es
Caride-Castro
Amado
A
Martinez-Zorzano
Vicenta S
VS
De La Cadena
María Páez
MP
Rodríguez-Berrocal
Francisco J
FJ
D000230:Adenocarcinoma / Q000737:chemistry*; D014408:Biomarkers, Tumor / Q000032:analysis / Q000737:chemistry; D001706:Biopsy; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D015179:Colorectal Neoplasms / Q000737:chemistry*; D003208:Concanavalin A / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005643:Fucose / Q000032:analysis*; D006023:Glycoproteins / Q000032:analysis* / Q000737:chemistry*; D006801:Humans; D007413:Intestinal Mucosa / Q000737:chemistry*; D037102:Lectins / Q000302:isolation & purification* / Q000378:metabolism; D008567:Membranes, Artificial; D008970:Molecular Weight; D019158:N-Acetylneuraminic Acid / Q000032:analysis*; D009928:Organ Specificity
D014408:Biomarkers, Tumor; D006023:Glycoproteins; D037102:Lectins; D008567:Membranes, Artificial; D003208:Concanavalin A; D005643:Fucose; D019158:N-Acetylneuraminic Acid