Location of Disulfide bonds in mature α-L;-fucosidase from pea

✍ Scribed by Anna Codina; Marta Vilaseca; Teresa Tarragó; Irene Fernández; Dolors Ludevid; Ernest Giralt

• Publisher: John Wiley and Sons
• Year: 2001
• Tongue: English
• Weight: 663 KB
• Volume: 7
• Category: Article
• ISSN: 1075-2617
• DOI: 10.1002/psc.323

✦ Synopsis

Abstract

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.