In this work, peptides selected from a microarray were found to inhibit β-gal with promiscuous mechanisms. Peptides inhibited the enzyme in a noncompetitive kinetics, and the inhibition of enzyme activities was reduced under high enzyme concentrations and the addition of detergent. Dynamic light scattering and atomic force microscope revealed that peptide/enzyme aggregation was related to inhibited enzyme activities. Positively charged residues of arginine and lysine were critical for the enzyme inhibition. The preincubation of peptide inhibitors with negatively charged biopolymers of polyphosphates, ssDNA, and low pI peptides could increase the residual activity of peptide-inhibited enzyme, possibly due to the disruption of the electrostatic interaction between positively charged peptide residues and the β-gal surface. Further, negative biopolymers were able to recover the activity of the aggregated peptide/β-gal complex. Negatively charged biopolymers could be used in high-throughput screening assays to reduce peptides/protein aggregation and thereby minimize promiscuous inhibitions.
Keywords: activity recovery; biopolymers; enzyme aggregation; peptide promiscuous inhibitor; β-galactosidase.