Abstract
A keratinase from Pseudomonas aeruginosa (KerPA), which belongs to the M4 family of metallopeptidases, was characterised in this study. This enzyme was engineered with non-canonical amino acids (ncAAs) using genetic code expansion. Several variants with enhanced activity and thermostability were identified and the most prominent, Y21pBpF/Y70pBpF/Y114pBpF, showed an increase in enzyme activity and half-life of approximately 1.3-fold and 8.2-fold, respectively. Considering that keratinases usually require reducing agents to efficiently degrade keratin, the Y21pBpF/Y70pBpF/Y114pBpF variant with enhanced activity and stability under reducing conditions may have great significance for practical applications. Molecular Dynamics (MD) was performed to identify the potential mechanisms underlying these improvements. The results showed that mutation with pBpF at specific sites of the enzyme could fill voids, form new interactions, and reshape the local structure of the active site of the enzyme.
Highlights
The increasing consumption of chicken has led to the yearly production of millions of tons of feathers, which are the main by-products of poultry production (Li, 2019)
The crystal structure of the solvent tolerant elastase from P. aeruginosa strain K was used as a template (PDB ID: 4k89.1) to generate the structural model of keratinase from Pseudomonas aeruginosa (KerPA) using the SWISS-MODEL server (Figure 1C)
Keratinases that can degrade keratin-rich waste in an environmentally friendly manner have been the focus of increasing attention
Summary
The increasing consumption of chicken has led to the yearly production of millions of tons of feathers, which are the main by-products of poultry production (Li, 2019). The disposal of the vast amounts of feathers produced directly to the environment would be far beyond the capacity of nature to decompose them in a timely manner and may facilitate the growth and outbreaks of various pathogenic microorganisms. This manner of feather disposal could seriously threaten both economic animal and human health (Cavello et al, 2012), whereas burning feather waste is not encouraged because it causes air pollution.
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