Inhibition of Granzyme A (unknown origin) expressed in Pichia pastoris assessed as second- order inhibition rate constant (Kobs/I ratio) pre-incubated with SK15 substrate for 15 mins before enzyme addition by spectrofluorometry
Binding affinity to Granzyme A (unknown origin) expressed in Pichia pastoris assessed as appearance of fluorescent 35 kDa species at enzyme to probe ratio was 1:1 incubated for 5 mins by SDS-PAGE based in-gel detection method
Binding affinity to Granzyme A (unknown origin) expressed in Pichia pastoris assessed as minimum concentration needed for GrA labeling at enzyme to probe ratio was 1:1 incubated for 5 to 30 mins by SDS-PAGE based in-gel detection method
Binding affinity to Granzyme A active form (unknown origin) expressed in Pichia pastoris at 500 nM in presence of 1 uM methyl 4-[(3S)-3-[[2-[(2S)-2-[(1-diphenoxyphosphoryl-4-guanidinobutyl)carbamoyl]-2,3,3a,4,5,6,7,7a-octahydroindol-1-yl]-2-oxoethyl]carbamoyl]-3,4-dihydro-1H-isoquinolin-2-yl]-4-oxobutanoate by Western blot analysis
Binding affinity to Granzyme A zymogen form (unknown origin) expressed in Pichia pastoris at 500 nM in presence of 1 uM compound 3 by Western blot analysis
Binding affinity to Granzyme A in human NK92 cells at 1 uM treated 1 hr after GrA covalent inhibitor methyl 4-[(3S)-3-[[2-[(2S)-2-[(1-diphenoxyphosphoryl-4-guanidinobutyl)carbamoyl]-2,3,3a,4,5,6,7,7a-octahydroindol-1-yl]-2-oxoethyl]carbamoyl]-3,4-dihydro-1H-isoquinolin-2-yl]-4-oxobutanoate dosing and measured 30 mins to 2 hrs post dose by SDS-PAGE analysis