Enzymatic activity and stability can be increased by encapsulation in ribonucleoprotein nanoparticle cages, called vaults. Vault nanocages are found in many eukaryotic organisms, and can be engineered to package specific macromolecules. In this study, a ligninolytic enzyme, manganese peroxidase (MnP), which biodegrades a broad range of organic contaminants, was used as a model enzyme. We genetically attached a zip-code domain, called INT, to synthesize enzyme-INT fusion, MnP-INT, thereby ensuring strong selective packaging inside the vault nanoparticles. Both MnP-INT and vault-packaged MnP-INT showed viable peroxidase activity, and successfully degraded munition constituents, which are carcinogens and environmentally persistent pollutants, as well as bisphenols (BPs), which are endocrine-disrupters and reproductive toxicants. Further stability tests confirmed that packaging into vaults significantly enhanced MnP-INT longevity by inhibiting the second enzyme deactivation step. Vault-packaged MnP (vMnP) with an activity of 150 U/L catalyzed 72% removal of 2-amino-4-nitrotoluene (ANT), and 40 U/L vMnP catalyzed 59% removal of 2,4-diaminotoluene(DAT). At lower enzyme activity of vMnP, significant biodegradation of ANT and DAT was observed whereas natural MnP (nMnP) isolated from wood-decaying fungus, <i>Phanerochaete chrysosporium</i>, showed insignificant removal. Over a 24-hour period, vMnP treatment removed 80-95% BPA, BPF, and BPAP at lower enzyme dosage, while nMnP only resulted in a 19-36% removal. Further toxicity studies in <i>Caenorhabditis elegans </i>revealed that vMnP treatment completely diminished BPs’ adverse effects on germline apoptosis and fertility, which are the hallmarks of reproductive toxicity, but nMnP treatment did not. Collectively, these results indicate that fungal enzymes packaged in vault nanoparticles are more stable and more efficient in removing and detoxifying contaminants. Vault packaging presents an effective enzymatic biodegradation technology for a variety of compounds in industrial and environmental applications.

Vault packaged ligninolytic enzymes as nanoscale environmental bioremediation agents

2019 Spring National Meeting

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