PDBsum entry 3g01

Hydrolase

PDB id: 3g01

Contents

Protein chains

211 a.a. *

Waters ×28

* Residue conservation analysis

PDB id:

3g01

Name:

Hydrolase

Title:

Structure of grc mutant e192r/e193g

Structure:

GranzymE C. Chain: a, b. Fragment: unp residues 21-247. Synonym: cytotoxic cell protease 2, ccp2, b10. Engineered: yes. Mutation: yes

Source:

Mus musculus. Mouse. Organism_taxid: 10090. Expressed in: pichia pastoris. Expression_system_taxid: 4922.

Resolution:

2.50Å R-factor: 0.213 R-free: 0.236

Authors:

A.M.Buckle,D.Kaiserman,J.C.Whisstock

Key ref:

D.Kaiserman et al. (2009). Structure of granzyme C reveals an unusual mechanism of protease autoinhibition. Proc Natl Acad Sci U S A, 106, 5587-5592. PubMed id: 19299505 DOI: 10.1073/pnas.0811968106

Date:

27-Jan-09 Release date: 17-Mar-09

Protein chains

P08882  (GRAC_MOUSE) -  Granzyme C from Mus musculus

Seq: 248 a.a.

Struc: 211 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.4.21.- - ?????

DOI no: 10.1073/pnas.0811968106

Proc Natl Acad Sci U S A 106:5587-5592 (2009)

PubMed id: 19299505

Structure of granzyme C reveals an unusual mechanism of protease autoinhibition.

D.Kaiserman, A.M.Buckle, P.Van Damme, J.A.Irving, R.H.Law, A.Y.Matthews, T.Bashtannyk-Puhalovich, C.Langendorf, P.Thompson, J.Vandekerckhove, K.Gevaert, J.C.Whisstock, P.I.Bird.

ABSTRACT

Proteases act in important homeostatic pathways and are tightly regulated. Here, we report an unusual structural mechanism of regulation observed by the 2.5-A X-ray crystal structure of the serine protease, granzyme C. Although the active-site triad residues adopt canonical conformations, the oxyanion hole is improperly formed, and access to the primary specificity (S1) pocket is blocked through a reversible rearrangement involving Phe-191. Specifically, a register shift in the 190-strand preceding the active-site serine leads to Phe-191 filling the S1 pocket. Mutation of a unique Glu-Glu motif at positions 192-193 unlocks the enzyme, which displays chymase activity, and proteomic analysis confirms that activity of the wild-type protease can be released through interactions with an appropriate substrate. The 2.5-A structure of the unlocked enzyme reveals unprecedented flexibility in the 190-strand preceding the active-site serine that results in Phe-191 vacating the S1 pocket. Overall, these observations describe a broadly applicable mechanism of protease regulation that cannot be predicted by template-based modeling or bioinformatic approaches alone.

Selected figure(s)

Figure 2.
Granzyme C inactivation is caused by a register shift. (A) Superposition of granzyme C (cyan) with human granzyme A (yellow; PDB ID code 1ORF). The active-site triads are shown as stick models: magenta, granzyme C; green, granzyme A. The blue (granzyme C) and red (granzyme A) regions are enlarged in B. (B) Stereoview of the register shift in granzyme C. The arrow indicates the direction of register shift. (C) Stereoview of the oxyanion hole. Trypsin in complex with BPTI (PDB ID code 2PTC) was added to the superposition. BPTI residues 13–17 indicate likely positioning of substrate P3–P2′. Dashed lines indicate hydrogen bonds between granzyme C E193 carbonyl and S195 amide and Oγ. The figures were rendered with PyMOL (32).

Figure 4.
Stereochemistry of Glu-192. A stereo image of the F[o] − F[c] omit map, contoured at 3.0 σ, generated after coordinate refinement in the absence of residues Phe-191, Glu-192, and Glu-193. These residues appear as stick models; the salt bridge with Arg-99 is also shown for clarity.

Figures were selected by an automated process.