510-0005 g-linked ATP agarose resin

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Properties for g-linked ATP agarose resin

Product Category Others
Quantity 5 ml
Shipping to Not USA/Canada
PDF datasheet View Datasheet
Manufacturer Novus Biologicals Inc.

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Affinity resins have been widely used for the purification of enzymes that bind nucleotides and related molecules. Resins in which ATP is linked via the g-phosphate have been valuable in identifying proteins in the purine-binding proteome, which includes kinases, heat shock proteins and other ATP-binding proteins.

KinaseBind(TM) resin comprises ATP attached to agarose beads via its g-phosphate. Two forms of the resin are available with low and high ligand substitution. A long hydrophilic spacer (14-atom) is used to minimise unwanted hydrophobic interactions and to facilitate unhindered interactions with biomolecules.


2.1. Storage of KinaseBind(TM) resin
The resin is supplied as 50% (v/v) slurry in 10mM Tris/300 mM NaCl/1mM EDTA, pH 8.0. The product is shipped at ambient temperature but should be stored at 4C.

2.2. Materials required (but not supplied)
For small sample volumes you may need only a microfuge and 1.5 ml tubes. For larger volumes (up to 20 ml) the purification of binding proteins is conveniently carried out using disposable polypropylene columns. A simple mixing device (e.g. rotary shaker or end-over-end mixer) may also be useful.

2.3. Overview of procedure
KinaseBind(TM) resin is added to a crude protein extract and the suspension is gently mixed. After a period of incubation the resin is transferred to a disposable column and washed to remove non-bound or loosely adsorbed material. Finally, the column is eluted with buffer containing a competing ligand.

Since KinaseBind(TM) resin may be used to capture many purine-binding proteins the instructions below provide only general guidance on the use of resin. You may need to modify the conditions to facilitate the binding of your particular biomolecule of interest.

2.4 Buffers
For simplicity we would recommend that you start with the same buffer for the equilibration, binding and wash steps. The elution buffer is prepared by adding a competing ligand.

2.4.1 Types of buffers
The buffer and pH must be compatible with the biomolecule of interest. Tris (pH 7.5-8.5) and Hepes (pH 7.0-8.0) are commonly used but other buffers may also be suitable.

2.4.2 Metal ions
ATP-binding proteins usually recognise ATPmagnesium ion complexes rather than free ATP. It is usual to include MgCl2 (at least 10 mM) in all column buffers to facilitate metal-dependent interactions with the resin.

2.4.3 Salts
To prevent non-specific electrostatic interactions with the matrix include 100mM


ATP linked to a support via its g-phosphate has been used for affinity purification of kinases, heat-shock proteins and other ATP-binding proteins.

KinaseBind is a g-phosphate-linked ATP resin with a long hydrophillic spacer that facilitates molecular interactions with biomolecules. Linkage of ATP via the terminal phosphate means that the immobilised ligand cannot be removed by phosphatases in crude extracts.

Storage Store at 4C. Do not freeze.
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