Protein tethering kit (cysteine)

$818

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This kit applies cysteine–maleimide chemistry to attach DNA handles to your protein of interest for subsequent optical-tweezers measurements of protein folding and conformational changes. We recommend this labeling procedure over ybbR labeling, but it requires the absence of native cysteine residues on the protein of interest, especially in the functional domain of the protein.

For labeling, short maleimide-modified oligonucleotides react with specifically incorporated cysteines in the protein. These oligonucleotides then anneal with the complementary 5’-overhangs of two DNA handles.

The kit also includes purified and functionalized adenosine kinase (AdK) that can be used as a positive control and quality assurance for each step of the labeling procedure.

Our Protein tethering kit relieves you from costs and time that are typically required for personnel training, troubleshooting, experiment optimization, and custom application development. After finishing this kit, new materials can easily be ordered as a recurring package.

Material supplied:

  • Maleimide-modified oligos
  • Biotin- and digoxigenin-modified DNA handles (520 bp per handle)
  • Reducing agent TCEP
  • 6xHis tagged AdK protein
  • Buffer 40 mM Tris HCl, 50 mM NaCl
  • Streptavidin coated polystyrene beads (1.76 µm)
  • Anti-digoxigenin coated polystyrene beads (0.75 µm)
  • Estimated for 2 weeks of measurements (handles and beads, more can be ordered as Protein handles kit (recurring))
  • Estimated for 8 weeks of measurements (labeled protein and control)
  • Estimated 1-3 weeks delivery time

Specifications

Maleimide-modified oligonucleotides:

Dry pellet (not visible), 2 vials. Lyophilized oligonucleotides for protein labeling using maleimide-cysteine chemistry. Store at -20°C.

Biotinylated and digoxigenin-labeled DNA handles (529 bp):

4 μl, 10 vials. Handles mix (50/50) with an overhang complementing the maleimide-modified oligonucleotides. Store all materials at -80°C.

TCEP (10 mM):

5 μl, 2 vials | 10 mM. Reducing agent. Store all materials at -80°C.

Customized protein:

40 μl | 0.5 μg/μl (20 μM). Store all materials at -80°C.

Streptavidin-coated silica beads (⌀ 1.0–1.4 μm):

25 μl | 1% (w/v). Beads in PBS with 3 mM sodium azide, with a specific diameter (e.g., 1.2 μm) within the given range. Store all materials at +4°C.

Anti-digoxigenin-coated polystyrene beads (⌀ 0.7–0.9 μm):

60 μl | 0.1% (w/v). Beads in PBS with 3 mM sodium azide, with a specific diameter (e.g., 0.8 μm) within the given range. Store all materials at +4°C.

Download the Full Specifications  or View the Experimental Protocols

Product Resources

Want to learn more?

See our Protein Folding and Conformational Changes application notes.

Want to use this product in practice?

Find out more about the C-Trap here.