BLIP-II Employs Differential Hotspot Residues To Bind Structurally Similar Staphylococcus aureus PBP2a and Class A ?-Lactamases.

ABSTRACT: The interaction of ?-lactamase inhibitory protein II (BLIP-II) with ?-lactamases serves as a model system to investigate the principles underlying protein-protein interactions. Previous studies have focused on identifying the determinants of binding affinity and specificity between BLIP-II and class A ?-lactamases. However, interactions between BLIP-II and other bacterial proteins have yet to be explored. Here, we provide evidence that BLIP-II binds penicillin binding protein 2a (PBP2a) from methicillin-resistant Staphylococcus aureus (MRSA) with a KD in the low micromolar range. In comparison to the binding constants for the potent interaction between BLIP-II and TEM-1 ?-lactamase (KD = 0.5 pM), the on-rate for BLIP-II binding PBP2a is 44?000 times slower and the off-rate is 170 times faster. Therefore, a slow association rate is a limiting factor for the potency of the interaction between BLIP-II and PBP2a. Results from alanine scanning mutagenesis of the predicted interface residues of BLIP-II indicate that charged residues on the periphery of the BLIP-II interface play a critical role for binding PBP2a, in contrast to previous findings that aromatic residues at the center of the BLIP-II interface are critical for the interaction with ?-lactamases. Interestingly, many of the alanine mutants at the BLIP-II interface increase kon for binding PBP2a, consistent with the association rate being a limiting factor for affinity. In summary, the results of the study reveal that BLIP-II binds PBP2a, although weakly compared to binding of ?-lactamases, and provides insights into the different binding strategies used for these targets.

SUBMITTER: Adamski CJ 

PROVIDER: S-EPMC5641629 | biostudies-literature | 2017 Feb

REPOSITORIES: biostudies-literature