2F9N: Crystal Structure Of The Recombinant Human Alpha I Tryptase Mutant K192qD216G IN COMPLEX WITH LEUPEPTIN

Tryptases alpha and beta are trypsin-like serine proteinases expressed in large amounts by mast cells. Beta-tryptase is a tetramer that has enzymatic activity, but requires heparin binding to maintain functional and structural stability, whereas alpha-tryptase has little, if any, enzymatic activity but is a stable tetramer in the absence of heparin. As shown previously, these differences can be mainly attributed to the different conformations of the 214-220 segment. Interestingly, the replacement of Asp216 by Gly, which is present in beta-tryptase, results in enzymatically active but less stable alpha-tryptase mutants. We have solved the crystal structures of both the single (D216G) and the double (K192Q/D216G) mutant forms of recombinant human alphaI-tryptase in complex with the peptide inhibitor leupeptin, as well as the structure of the non-inhibited single mutant. The inhibited mutants exhibited an open functional substrate binding site, while in the absence of an inhibitor, the open (beta-tryptase-like) and the closed (alpha-tryptase-like) conformations were present simultaneously. This shows that both forms are in a two-state equilibrium, which is influenced by the residues in the vicinity of the active site and by inhibitor/substrate binding. Novel insights regarding the observed stability differences as well as a potential proteolytic activity of wild-type alpha-tryptase, which may possess a cryptic active site, are discussed.
PDB ID: 2F9NDownload
MMDB ID: 102599
PDB Deposition Date: 2005/12/6
Updated in MMDB: 2012/09
Experimental Method:
x-ray diffraction
Resolution: 1.6  Å
Source Organism:
synthetic construct
Similar Structures:
Biological Unit for 2F9N: octameric; determined by author and by software (PISA)
Molecular Components in 2F9N
Label Count Molecule
Proteins (8 molecules)
Alpha I Tryptase
Molecule annotation
Molecule annotation
Chemicals (14 molecules)
* Click molecule labels to explore molecular sequence information.

Citing MMDB