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Location of Disulfide bonds in mature α‐ L ;‐fucosidase from pea
Author(s) -
Codina Anna,
Vilaseca Marta,
Tarragó Teresa,
Fernández Irene,
Ludevid Dolors,
Giralt Ernest
Publication year - 2001
Publication title -
journal of peptide science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 66
eISSN - 1099-1387
pISSN - 1075-2617
DOI - 10.1002/psc.323
Subject(s) - chemistry , residue (chemistry) , acrylamide , disulfide bond , alkylation , cleavage (geology) , disulfide linkage , peptide , posttranslational modification , stereochemistry , enzyme , biochemistry , combinatorial chemistry , cysteine , organic chemistry , catalysis , geotechnical engineering , fracture (geology) , engineering , copolymer , polymer
Fuc‐9 is the mature form of a vacuolar α‐ L ;‐fucosidase enzyme which seems to play an important role in plant growth regulation. Fuc‐9 is a 202‐residue protein containing five Cys residues located at positions 64, 109, 127, 162 and 169. In this study, the disulfide structure of Fuc‐9 was determined by MALDI‐TOF mass spectrometry (MS), with minimal clean‐up of the samples and at a nanomolar scale. Two strategies, based on a specific chemical cleavage (with 2‐nitro‐5‐thiocyanobenzoic acid and alkaline conditions) at the Cys residues and modification of Cys residues by acrylamide/deuterium labeled acrylamide alkylation, were used. Using these methods, the disulfide pairings Cys64‐Cys109 and Cys162‐Cys169 could be established. The advantages and limitations of our experimental approach are discussed. Copyright © 2001 European Peptide Society and John Wiley & Sons, Ltd.