The process of recycling poly(ethylene terephthalate) (PET) remains a major challenge due to the enzymatic degradation of high-crystallinity PET (hcPET). Recently, a bacterial PET-degrading enzyme, PETase, was found to have the ability to degrade the hcPET, but with low enzymatic activity. Here we present an engineered whole-cell biocatalyst to simulate both the adsorption and degradation steps in the enzymatic degradation process of PETase to achieve the efficient degradation of hcPET. Our data shows that the adhesive unit hydrophobin and degradation unit PETase are functionally displayed on the surface of yeast cells. The turnover rate of the whole-cell biocatalyst toward hcPET (crystallinity of 45%) dramatically increases approximately 328.8-fold compared with that of purified PETase at 30 °C. In addition, molecular dynamics simulations explain how the enhanced adhesion can promote the enzymatic degradation of PET. This study demonstrates engineering the whole-cell catalyst is an efficient strategy for biodegradation of PET.

Significance: Amyotrophic lateral sclerosis (ALS) is an invariably fatal neurodegenerative disease. Autosomal dominant mutations in the SOD1 gene are responsible for 12% of familial ALS cases and 1.5% of sporadic cases. However, it remains unknown whether the large fibrillar aggregates formed by misfolded SOD1 are a causative agent in disease progression. By designing mutations that specifically stabilize SOD1 fibrils or nonnative oligomers, we found that the assembly into insoluble fibrils mitigated the neurotoxic effects caused by aberrant conformation of trimers. We also demonstrated with electron microscopy that toxic SOD1 trimers displayed heterogeneous structures, in concert with computational studies. Our findings suggest a protective role of fibrils and a plausible pharmaceutical strategy, as promoting SOD1 fibrillogenesis reduces the population of neurotoxic species.

CRISPR/Cas12a systems have been repurposed as powerful tools for developing next-generation molecular diagnostics due to their trans-cleavage ability. However, it was long considered that the CRISPR/Cas12a system could only recognize DNA targets. Herein, we systematically investigated the intrinsic trans-cleavage activity of the CRISPR/Cas12a system (LbCas12a) and found that it could be activated through fragmented ssDNA activators. Remarkably, we discovered that the single-stranded DNA (ssDNA) activators in the complementary crRNA-distal domain could be replaced by target miRNA sequences without the need for pre-amplification or specialized recognition mechanisms. Based on these findings, we proposed the “Fragment Complementary Activation Strategy” (FCAS) and designed reverse fluorescence-enhanced lateral flow test strips (rFLTS) for the direct detection of miRNA-10b, achieving a limit of detection (LOD) of 5.53 fM and quantifying the miRNA-10b biomarker in clinical serum samples from glioma patients. Moreover, for the first time, we have developed the FCAS-based CRISPR/Cas12a system for miRNA in situ imaging, effectively recognizing tumor cells. The FCAS not only broadens the scope of CRISPR/Cas12a system target identification but also unlocks the potential for in-depth studies of CRISPR technology in many diagnostic settings.