HRV 3C Protease, 1mg (1ug/1u)

HRV 3C Protease

Description

Human rhinovirus 3C protease (HRV3C Protease) is a cysteine protease that recognizes the cleavage site of Leu-Glu-Val-Leu-Phe-Gln/Gly-Pro, commonly referred as the PreScission Site. It cleaves between Gln and Gly. The recombinant form of the HRV3C protease is a restriction grade protease that has robust activity at 4oC and high specific activity. This HRV3C Protease is a 47 kDa protein with both GST and His tags so it can be removed by either Ni chelating or Glutathione (GSH) resin.

Activity and Specificity

The activity of HRV3C Protease is tested using a control cleavage protein. One ?g of HRV3C Protease has an equivalent of ~1 units conventionally used by other suppliers (1 unit of HRV3C protease cleaves >95% target cleavage protein at 4oC for 16 hours.).

A GST-fusion protein (~49 kDa) at 1 mg/ml is incubated with HRV3C at a ratio of (1) 1:100, (2) 1:200, (3) 1:400 (w/w) in a buffer of 25 mM Tris-HCl, pH7.5, 150 mM NaCl, 5 mM DTT at 4oC for 16 hours. The cleaved products are 27 kDa and 22 kDa. A, HRV3C protease; B and C, competitor HRV3C protease.


Component

HRV3C Protease 1 mg/500 ?l per tube at 2 mg/ml in
50 mM Tris-HCl, pH8.0
150 mM NaCl
1 mM EDTA
1 mM TCEP
50% glycerol

Storage

Store HRV3C Protease at -20oC.

Cleavage Condition

It is recommended to test the HRV3C Protease cleavage with protease-to-target protein ratio of 1:100 (w/w) in the buffer suitable for the target protein at 4oC overnight, with the target protein concentration at or below 1 mg/ml. Under this condition, >95% of target protein is cleaved in most cases. The efficiency of cleavage varies due to the sequences around the cleavage site, the conformation and the solubility of the target protein. Due to its high specificity, more protease can be used without non-specific cleavage of target proteins. No non-specific cleavage of control cleavage protein is observed at protease-to-control cleavage protein ratio of 1:10.

Removal of HRV3C Protease after Cleavage

The HRV3C Protease contains both GST and His tags. After cleavage of the target protein, HRV3C Protease can be easily removed along with the tags from the cleavage reaction by affinity chromatography on a Ni chelating resin for His-tagged target protein or GSH resin for GST-tagged target protein.