Investigation of the Active Center of Porcine‐Pancreatic Amylase

The effect of maltose was studied in porcine pancreatic amylase. At neutral pH 1% (29 mM) maltose produced with amylase a difference spectrum characteristic of the perturbation of tryptophan. The molar absorption difference at the maximum wavelength was Δ 290 = 1200. The difference spectrum appeared to be specific for maltose. Perturbation difference spectra measurements in 20% polyethylene glycol indicated that one tryptophyl side chain per mol amylase was involved in the interaction with maltose. The dissociation constant of the amylase · maltose complex calculated from the concentration dependence of the absorption difference at 290 nm was K s = 13 mM. Maltose inhibited amylase activity competitively and an inhibition constant of K i = 25 mM was obtained, a similar value to that found spectrophotometrically. It is assumed that the tryptophyl side chain interacting with maltose may be involved in the binding of substrate by pancreatic amylase.