American Chemical Society
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Rapid Profiling of Peptide Stability in Proteolytic Environments

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journal contribution
posted on 2009-02-15, 00:00 authored by Hans H. Gorris, Steffen Bade, Niels Röckendorf, Eike Albers, M. Alexander Schmidt, Milan Fránek, Andreas Frey
The notorious degradation susceptibility of peptides is a major obstacle to their use as medicinal drugs. Assays with which the stability of peptides in complex proteolytic environments can be determined are thus indispensable for peptide drug development. Herein, we describe a new peptide proteolysis assay that meets that demand. It unites the high-throughput capacity of heterogeneous with the well-defined kinetic characteristics of homogeneous assay formats and operates on the cleavage-caused loss of a detection handle. We have confirmed the assay’s accuracy with well-defined model interactions and proved its high versatility and robustness with a representative application where we determined the half-lives of 375 different peptides in a crude intestinal protease preparation. With this reliable, reproducible, and efficient assay the enzyme kinetics of any peptide−protease interaction is accessible even for complex protease solutions.