Cystatin inhibition of cathepsin B requires dislocation of the proteinase occluding loop. Demonstration By release of loop anchoring through mutation of his110.
Cystatins A and C were both shown to inhibit cathepsin B by a two-step mechanism, involving an initial weak interaction followed by a conformational change. Disruption of the major salt bridge anchoring the occluding loop of cathepsin B to the main body of the enzyme by mutation of His110 to Ala converted the binding to an apparent one-step reaction. The second step of cystatin binding to cathepsin B must therefore be due to the inhibitor having to alter the conformation of the enzyme by displacing the occluding loop to allow a tight complex to be formed. Cystatin A was appreciably less effective in displacing the loop than cystatin C, resulting in a considerably lower overall inhibition rate constant.
Pavlova
A
A
The Biomedical Center, Department of Veterinary Medical Chemistry, Swedish University of Agricultural Sciences, Uppsala, Sweden.
Krupa
J C
JC
Mort
J S
JS
Abrahamson
M
M
Björk
I
I
D000409:Alanine; D019943:Amino Acid Substitution; D002401:Cathepsin B / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000235:genetics; D055316:Cystatin C; D015891:Cystatins / Q000494:pharmacology*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D006639:Histidine; D006801:Humans; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000494:pharmacology
C525030:CST3 protein, human; D055316:Cystatin C; D015891:Cystatins; D015853:Cysteine Proteinase Inhibitors; D011994:Recombinant Proteins; D006639:Histidine; D002401:Cathepsin B; D000409:Alanine