Structure of granzyme C reveals an unusual mechanism of protease autoinhibition.
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.
Kaiserman
Dion
D
Department of Biochemistry and Molecular Biology, Monash University, Clayton 3800, Australia.
Buckle
Ashley M
AM
Van Damme
Petra
P
Irving
James A
JA
Law
Ruby H P
RH
Matthews
Antony Y
AY
Bashtannyk-Puhalovich
Tanya
T
Langendorf
Chris
C
Thompson
Philip
P
Vandekerckhove
Joël
J
Gevaert
Kris
K
Whisstock
James C
JC
Bird
Phillip I
PI
D000596:Amino Acids; D000818:Animals; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D053804:Granzymes / Q000737:chemistry*; D011487:Protein Conformation; D012377:Rodentia; D012697:Serine Endopeptidases / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors
D000596:Amino Acids; D015842:Serine Proteinase Inhibitors; D053804:Granzymes; D012697:Serine Endopeptidases