title: Identification of peptide inhibitors of penicillinase using a phage display library.
abstract: There is a constant need to identify novel inhibitors to combat β-lactamase-mediated antibiotic resistance. In this study, we identify three penicillinase-binding peptides, P1 (DHIHRSYRGEFD), P2 (NIYTTPWGSNWS), and P3 (SHSLPASADLRR), using a phage display library. Surface plasmon resonance (SPR) is utilized for quantitative determination and comparison of the binding specificity of selected peptides to penicillinase. An SPR biosensor functionalized with P3-GGGC (SHSLPASADLRRGGGC) is developed for detection of penicillinase with excellent sensitivity (15.8 RU nM(-1)) and binding affinity (KD = 0.56 nM). To determine if peptides can be good inhibitors for penicillinase, these peptides are mixed with penicillinase and their inhibition efficiency is determined by measuring the hydrolysis of substrate penicillin G using UV-vis spectrophotometry. Peptide P2 (NIYTTPWGSNWS) is found to be a promising penicillinase inhibitor with a Ki of 9.22 μM and a Ki' of 33.12 μM, suggesting that the inhibition mechanism is a mixed pattern. This peptide inhibitor (P2) can be used as a lead compound to identify more potent small molecule inhibitors for penicillinase. This study offers a potential approach to both detection of β-lactamases and development of novel inhibitors of β-lactamases.
authors.affiliation: Department of Chemical and Biomolecular Engineering, National University of Singapore, 4 Engineering Drive 4, Singapore, 117585, Singapore.
authors.forename: Qiongjing
authors.initials: Q
authors.lastname: Zou
authors.affiliation: Department of Chemical and Biomolecular Engineering, National University of Singapore, 4 Engineering Drive 4, Singapore, 117585, Singapore. Electronic address: cheyk@nus.edu.sg.
authors.forename: Kun-Lin
authors.initials: KL
authors.lastname: Yang
mesh_terms: D000595:Amino Acid Sequence; D006868:Hydrolysis; D007700:Kinetics; D010400:Penicillin G / Q000378:metabolism; D010405:Penicillinase / Q000737:chemistry / Q000378:metabolism*; D019151:Peptide Library; D011485:Protein Binding; D013056:Spectrophotometry, Ultraviolet; D013379:Substrate Specificity; D020349:Surface Plasmon Resonance; D065093:beta-Lactamase Inhibitors / Q000737:chemistry / Q000378:metabolism*
chemical_list: D019151:Peptide Library; D065093:beta-Lactamase Inhibitors; D010405:Penicillinase; D010400:Penicillin G
keywords: Antibiotic resistance; Enzyme inhibitor; Penicillinase; Peptide inhibitor; Phage display; Surface plasmon resonance