(23) Assay Card

General Information
Summary Both SFTI-1 and acyclic SFTI-1 are potent inhibitors of bovine beta trypsin
Condition Trypsin activity was assayed at 25¬C in a 96-well plate with 350 ul total reaction volume per well, with 42 nM bovine beta-trypsin (Type XIII TPCK treated, EC using BAPNA substrate (4.0 mM) and buffer (pH 8) (50 mM Tris-HCl, 25 mM CaCl2). SFTI-1 inhibitors (0-400 nM) and trypsin were incubated together in buffer for two hours at 25¬C. The reaction at 25¬C was initiated by addition of substrate and monitored at 405 nm with readings taken every 60 seconds for 120 minutes using a Molecular Devices SpectraMax 250 or a Labsystems Multiskan Ascent.
Result Cyclic SFTI-1 inhibited bovine beta-trypsin with a Ki of 0.5(±0.1) nM and acyclic SFTI also inhibited inhibited trypsin and had a Ki of 12.1(±3.2) nM.
AssayType Protease inhibition

Korsinczky ML, Schirra HJ, Rosengren KJ, West J, Condie BA, Otvos L, Anderson MA, Craik DJ (2001) Solution structures by 1H NMR of the novel cyclic trypsin inhibitor SFTI-1 from sunflower seeds and an acyclic permutant. J Mol Biol 311:579-91

Proteins Assayed N-SFTI-1-C