De novo design of alpha-amylase inhibitor: a small linear mimetic of macromolecular proteinaceous ligands.
We report a low molecular weight inhibitor of alpha-amylases based on a linear peptidic scaffold designed de novo through the use of combinatorial chemistry. The inhibitory motif denoted PAMI (peptide amylase inhibitor) was selected by using L-peptide libraries and was fine-tuned by the introduction of unnatural modifications. PAMI specifically inhibits glycoside hydrolases of family 13. Its interaction with porcine pancreatic alpha-amylase was characterized by inhibition kinetics, fluorescence competition assays with natural alpha-amylase inhibitors, and isothermal titration calorimetry. We demonstrate that the critical amino acid residues in PAMI are shared with those in the macromolecular proteinaceous inhibitors that, however, bind to alpha-amylases through a spatially scattered set of intermolecular contacts. Thus, natural molecular evolution as well as combinatorial evolution selected the same alpha-amylase binding determinants for completely different spatial frameworks.
Dolecková-Maresová
Lucie
L
Institute of Organic Chemistry and Biochemistry, Academy of Sciences of The Czech Republic, Flemingovo nam. 2, 16610 Praha, Czech Republic.
Pavlík
Manfred
M
Horn
Martin
M
Mares
Michael
M
D020816:Amino Acid Motifs; D000818:Animals; D001665:Binding Sites; D004791:Enzyme Inhibitors / Q000737:chemistry*; D007700:Kinetics; D008024:Ligands; D018716:Molecular Mimicry; D015394:Molecular Structure; D019151:Peptide Library; D013552:Swine; D000516:alpha-Amylases / Q000037:antagonists & inhibitors* / Q000737:chemistry*
D004791:Enzyme Inhibitors; D008024:Ligands; D019151:Peptide Library; D000516:alpha-Amylases