Biophysical characterization of a recombinant leucyl aminopeptidase from Bacillus kaustophilus

Abstract

The biophysical properties of Bacillus kaustophilus leucyl aminopeptidase (BkLAP) were examined in terms of analytical ultracentrifugation, fluorescence spectroscopy, and circular dichroism. By using the analytical ultracentrifuge, we demonstrated that tetrameric BkLAP exists as the major form in solution at protein concentration of 1.5 mg/ml at pH 8.0. The native enzyme started to unfold beyond ~1 M GdnHCl and reached an unfolded intermediate with [GdnHCl](1/2) at 1.8 M. Thermal unfolding of BkLAP was found to be highly irreversible and led to a marked formation of aggregates.

Comparison of Iron‐Binding Ability Between Thr70‐NapA and Ser70‐NapA of Helicobacter pylori

“…Then, the iron‐processed proteins were treated with different concentrations of guanidine hydrochloride (GdnHCl) at 30°C for 30 minutes. Fluorescence changes were measured at the excitation wavelength of 280 nm, and emission spectrum was scanned from 300 nm to 450 nm at room temperature .…”