Novel DNA-Binding Activity Exhibited by Poly(aspartic acid) Hydrolase‑1 Inhibits Poly(aspartic acid) Hydrolase Activity

Significant attention has been shifted toward the use and development of biodegradable polymeric materials to mitigate environmental accumulation and potential health impacts. One such material, poly­(aspartic acid) (PAA), is a biodegradable alternative to superabsorbent poly­(carboxylates), like poly­(acrylate). Three enzymes are known to hydrolyze PAA: PahZ1KT‑1 and PahZ2KT‑1 from Sphingomonas sp. KT-1 and PahZ1KP‑2 from Pedobacter sp. KP-2. We previously reported the X-ray crystal structure for PahZ1KT‑1, which revealed a homodimer complex with a strongly cationic surface spanning one side of each monomer. Here, we report the first characterization of any polymer hydrolase binding to DNA, where modeling data predict binding of the polyanionic DNA near the cationic substrate binding surface. Our data reveal that PahZ1 homologues from Sphingomonas sp. KT-1 and Pedobacter sp. KP-2 bind ssDNA and dsDNA with nanomolar binding affinities. PahZ1KT‑1 binds ssDNA and dsDNA with an apparent dissociation constant, K D,app = 81 ± 14 and 19 ± 1 nM, respectively, and these estimates are similar to the same behaviors exhibited by PahZ1KP‑2. Gel permeation chromatography data reveal that dsDNA binding promotes inhibition of PahZ1-catalyzed PAA biodegradation for each homologue. We propose a working model wherein binding of PahZ1 to extracellular biofilm DNA aids in the localization of the hydrolase to the environment in which PAA would first be encountered, thereby providing a mechanism to degrade extracellular PAA and potentially harvest aspartic acid for nutritional uptake.