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Peptide biotinylation with amine-reactive esters: differential side chain reactivity
B T Miller1, T J Collins, M E Rogers
1Department of Anatomy and Neurosciences, University of Texas Medical Branch, Galveston 77555-1043, USA.
Peptides
|January 1, 1997
Summary
Biotinylation reagents, commonly used for amino groups, can unexpectedly acylate other peptide functional groups. This study reveals novel biotinylation of arginine side chains in peptides, expanding understanding of these reactions.
Area of Science:
- Biochemistry
- Organic Chemistry
- Peptide Chemistry
Background:
- N-hydroxysuccinimide (NHS) esters of biotin are widely used for labeling proteins and peptides via reaction with amino groups.
- Previous understanding suggested specificity towards amino groups, but potential for side reactions was less explored.
Purpose of the Study:
- To investigate sequence-dependent acylation of peptides by various biotinylation reagents.
- To identify and characterize unexpected acylation sites beyond primary amino groups.
Main Methods:
- Utilized a model decapeptide, [D-Lys6]gonadotropin releasing hormone.
- Employed high-performance liquid chromatography (HPLC), amino acid analysis, hydroxylamine reaction, and mass spectrometry for product characterization.
Main Results:
- Confirmed O-acylation of serine (Ser4) and tyrosine (Tyr5) residues.
- Discovered a novel N-hydroxysuccinimide (NHS) ester biotinylation of the arginine (Arg8) side chain.
Conclusions:
- Commonly used biotinylation reagents exhibit broader reactivity than previously assumed.
- Sequence context significantly influences the acylation sites on peptides, including novel modifications of arginine residues.