NHS-biotin modification as a specific lysine probe coupled to mass spectrometry detection is increasingly used over the past years for assessing amino acid accessibility of proteins or complexes as an alternative when well-established methods are challenged1.
Numerous applications have already been reported2, with increasing use of N-HydroxySuccinimide- biotin (NHS-biotin) as a specific lysine probe3. As an example, determination of surface accessibility of amino acids may be useful to delineate protein–protein interfaces as shown in epitopemapping4.
Labeling of amino acids containing hydroxyl groups has been already observed for the Sulfo-NHS-biotin reagent reacting on model peptides33 and for 3,30-Dithiobis[sulfosuccinimidyl propionate] (DTSSP)5.It has been shown that serine, tyrosine, and threonine residues readily react with Sulfo- NHS-biotin if the hydroxyl-containing group is located two positions from a histidine residue6. It was proposed that the enhanced intrinsic reactivity of these hydroxyl groups was due to an interaction with the histidyl residue via hydrogen bonding that contributes to increase their nucleophilicity.
The pioneering observationsmade with DTSSP are also detected with NHS-biotin derivatives: hydroxyl groups of serine and tyrosine residues also reactwith primary amine reagents and serine label may be lost resulting in a dehydrated peptide. Use of biotin reagents allows affinity enrichment by means of streptavidin3, as well as absolute quantitation of the labels introduced per polypeptide by means of a spectrophotometric assay.
Use of the complete set of Sulfo-NHS-biotin reagents allows an efficient and reliable assignment of the different ions detected.
Reference:
1. G. Gabant, J. Augier et al, Assessment of solvent residues accessibility using three Sulfo-NHS-biotin reagents in parallel: application to footprint changes of a methyltransferase upon binding its substrate, J. Mass Spectrom. 2008; 43: 360–370
2. Glocker MO, Borchers C, Fiedler W, Suckau D, Przybylski M. Molecular characterization of surface topology in protein tertiary structures by amino-acylation and mass spectrometric peptide mapping. Bioconjugate Chem. 1994; 5: 583.
3. Azim-Zadeh O, Hillebrecht A, Linne U, Marahiel MA, Klebe G, Lingelbach K, Nyalwidhe J. Use of biotin derivatives to probe conformational changes in proteins. Journal of Biological Chemistry 2007; 282: 21609
4. Borch J, Jorgensen TJ, Roepstorff P. Mass spectrometric analysis of protein interactions. Current Opinion in Chemical Biology2005; 9: 509.
5. Swaim CL, Smith JB, Smith DL. Unexpected products from the reaction of the synthetic cross-linker 3, 3' dithiobis(sulfosuccinimidyl propionate), DTSSP with peptides. Journal of the American Society for Mass Spectrometry 2004; 15:736.
6. Miller BT, Kurosky A. Elevated intrinsic reactivity of seryl hydroxyl groups within the linear peptide triads His-Xaa-Ser or Ser-Xaa-His. Biochemical and Biophysical Research Communications 1993; 196: 461.