The ALFA-tag is a highly versatile tool for nanobody-based bioscience applications

Forskningsoutput: TidskriftsbidragArtikel i vetenskaplig tidskrift

Abstract

Specialized epitope tags are widely used for detecting, manipulating or purifying proteins, but often their versatility is limited. Here, we introduce the ALFA-tag, a rationally designed epitope tag that serves a remarkably broad spectrum of applications in life sciences while outperforming established tags like the HA-, FLAG®- or myc-tag. The ALFA-tag forms a small and stable α-helix that is functional irrespective of its position on the target protein in prokaryotic and eukaryotic hosts. We characterize a nanobody (NbALFA) binding ALFA-tagged proteins from native or fixed specimen with low picomolar affinity. It is ideally suited for super-resolution microscopy, immunoprecipitations and Western blotting, and also allows in vivo detection of proteins. We show the crystal structure of the complex that enabled us to design a nanobody mutant (NbALFAPE) that permits efficient one-step purifications of native ALFA-tagged proteins, complexes and even entire living cells using peptide elution under physiological conditions.

Detaljer

Författare
  • Hansjörg Götzke
  • Markus Kilisch
  • Markel Martínez-Carranza
  • Shama Sograte-Idrissi
  • Abirami Rajavel
  • Thomas Schlichthaerle
  • Niklas Engels
  • Ralf Jungmann
  • Pål Stenmark
  • Felipe Opazo
  • Steffen Frey
Enheter & grupper
Externa organisationer
  • Stockholms universitet
  • NanoTag Biotechnologies GmbH
  • University Medical Center Göttingen
  • Max Planck Institute of Biochemistry
  • Ludwig-Maximilian University of Munich
Forskningsområden

Ämnesklassifikation (UKÄ) – OBLIGATORISK

  • Medicinsk bioteknologi (med inriktning mot cellbiologi (inklusive stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci)
Originalspråkengelska
Artikelnummer4403
TidskriftNature Communications
Volym10
StatusPublished - 2019 sep 27
PublikationskategoriForskning
Peer review utfördJa