BioSITe: A Method for Direct Detection and Quantitation of Site-Specific Biotinylation

Dae In Kim, Jevon A. Cutler, Chan Hyun Na, Sina Reckel, Santosh Renuse, Anil K. Madugundu, Raiha Tahir, Hana L. Goldschmidt, Karen L. Reddy, Richard L. Huganir, Xinyan Wu, Natasha E. Zachara, Oliver Hantschel, Akhilesh Pandey

Research output: Contribution to journalArticle

16 Scopus citations

Abstract

Biotin-based labeling strategies are widely employed to study protein-protein interactions, subcellular proteomes and post-translational modifications, as well as, used in drug discovery. While the high affinity of streptavidin for biotin greatly facilitates the capture of biotinylated proteins, it still presents a challenge, as currently employed, for the recovery of biotinylated peptides. Here we describe a strategy designated Biotinylation Site Identification Technology (BioSITe) for the capture of biotinylated peptides for LC-MS/MS analyses. We demonstrate the utility of BioSITe when applied to proximity-dependent labeling methods, APEX and BioID, as well as biotin-based click chemistry strategies for identifying O-GlcNAc-modified sites. We demonstrate the use of isotopically labeled biotin for quantitative BioSITe experiments that simplify differential interactome analysis and obviate the need for metabolic labeling strategies such as SILAC. Our data also highlight the potential value of site-specific biotinylation in providing spatial and topological information about proteins and protein complexes. Overall, we anticipate that BioSITe will replace the conventional methods in studies where detection of biotinylation sites is important.

Original languageEnglish (US)
Pages (from-to)759-769
Number of pages11
JournalJournal of Proteome Research
Volume17
Issue number2
DOIs
StatePublished - Feb 2 2018
Externally publishedYes

Keywords

  • APEX
  • BioID
  • biotinylation
  • peptide
  • protein-protein interactions
  • proximity-dependent biotinylation
  • subcellular proteome

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

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  • Cite this

    Kim, D. I., Cutler, J. A., Na, C. H., Reckel, S., Renuse, S., Madugundu, A. K., Tahir, R., Goldschmidt, H. L., Reddy, K. L., Huganir, R. L., Wu, X., Zachara, N. E., Hantschel, O., & Pandey, A. (2018). BioSITe: A Method for Direct Detection and Quantitation of Site-Specific Biotinylation. Journal of Proteome Research, 17(2), 759-769. https://doi.org/10.1021/acs.jproteome.7b00775