Human rhinovirus 3C protease recognition site
Eight-residue site, LEVLFQGP, cleaved by human rhinovirus 3C protease between the glutamine and the glycine. Placed between a tag and its cargo, it lets the tag be removed after purification, leaving a short remnant on the cargo.
Origin: Human rhinovirus 3C protease recognition sequence
Characteristics
Encodes LEVLFQGP (8 amino acids, 24 nucleotides), with Glu at P5, Val at P4 and Gln-Gly-Pro across the scissile bond. Human rhinovirus 3C protease cuts between the glutamine and the glycine, so the released cargo keeps a Gly-Pro at its N terminus. For HRV-14 3C protease on synthetic peptides, TLFQ/GP is the minimal substrate, a P5 residue improves cleavage about 10-fold, and Val in place of the native Thr at P4 raises cleavage 4.8-fold. These peptides were HRV-14 sites, not LEVLFQGP itself.
Applications: Placed between an affinity or solubility tag and the protein of interest so that the tag can be removed with HRV 3C protease after capture, releasing the cargo with an N-terminal Gly-Pro. The cited literature maps the site requirements on synthetic peptides in vitro and does not report cleavage of a tagged fusion protein.
Limitations: Cleavage leaves Gly-Pro on the N terminus of the cargo and needs an added protease step. In HRV-14 3C protease peptide assays, Ala at P1 prime blocked cleavage and Ala at P2 prime lowered it about 10-fold, so the Gly-Pro after the Gln cannot be changed. Flanking sequence beyond the core changed cleavage of a 3A-3B site peptide 20-fold.
Sequence
References
- Cordingley et al. (1990). Substrate requirements of human rhinovirus 3C protease for peptide cleavage in vitro. J Biol Chem - Cordingley 1990 HRV 3C Substrate