Combined computational, rational, and empirical design of boiling-resistant keratinase.

ABSTRACT: Engineering of highly thermostable keratinase is of great theoretical interest in understanding protein stability mechanisms and practical significance for processing keratinous wastes such as feathers and wool. The thermostable subtilisin-like protease C2 is the major keratinase secreted by Thermoactinomyces vulgaris CDF but is rapidly inactivated at temperatures above 90°C. Here, we employed various methods to further stabilize protease C2. Four of the 35 selected single-point mutations designed by automated computational tools (PROSS, FireProt, ProteinMPNN, HyperMPNN, and ThermoMPNN) retained higher residual activity (~72%-84%) than protease C2 (~54%) after 1-h incubation at 85°C. The rational design of surface ion pairs and proline substitutions in β-turns generated two single-p

SUBMITTER: Yang Y 

PROVIDER: S-EPMC12838322 | biostudies-literature | 2026 Jan

REPOSITORIES: biostudies-literature