3MMG

Crystal structure of tobacco vein mottling virus protease

Summary for 3MMG

• Entry DOI: 10.2210/pdb3mmg/pdb
• Descriptor: Nuclear inclusion protein A, Nuclear inclusion protein B fragment, FORMIC ACID, ... (4 entities in total)
• Functional Keywords: 3c-type protease, tev, tvmv, viral protein, hydrolase
• Biological source: Tobacco vein mottling virus (TVMV); More
• Total number of polymer chains: 4
• Total formula weight: 56139.66

Authors

Ping, S.,Austin, B.P.,Tozser, J.,Waugh, D.S. (deposition date: 2010-04-19, release date: 2010-10-20, Last modification date: 2023-09-06)

Primary citation

Sun, P.,Austin, B.P.,Tozser, J.,Waugh, D.S.
Structural determinants of tobacco vein mottling virus protease substrate specificity.
Protein Sci., 19:2240-2251, 2010

PubMed Abstract: Tobacco vein mottling virus (TVMV) is a member of the Potyviridae, one of the largest families of plant viruses. The TVMV genome is translated into a single large polyprotein that is subsequently processed by three virally encoded proteases. Seven of the nine cleavage events are carried out by the NIa protease. Its homolog from the tobacco etch virus (TEV) is a widely used reagent for the removal of affinity tags from recombinant proteins. Although TVMV protease is a close relative of TEV protease, they exhibit distinct sequence specificities. We report here the crystal structure of a catalytically inactive mutant TVMV protease (K65A/K67A/C151A) in complex with a canonical peptide substrate (Ac-RETVRFQSD) at 1.7-Å resolution. As observed in several crystal structures of TEV protease, the C-terminus (∼20 residues) of TVMV protease is disordered. Unexpectedly, although deleting the disordered residues from TEV protease reduces its catalytic activity by ∼10-fold, an analogous truncation mutant of TVMV protease is significantly more active. Comparison of the structures of TEV and TVMV protease in complex with their respective canonical substrate peptides reveals that the S3 and S4 pockets are mainly responsible for the differing substrate specificities. The structure of TVMV protease suggests that it is less tolerant of variation at the P1' position than TEV protease. This conjecture was confirmed experimentally by determining kinetic parameters k(cat) and K(m) for a series of oligopeptide substrates. Also, as predicted by the cocrystal structure, we confirm that substitutions in the P6 position are more readily tolerated by TVMV than TEV protease.

PubMed: 20862670
DOI: 10.1002/pro.506

Experimental method

X-RAY DIFFRACTION (1.7 Å)