A method to site-specifically couple multiple enzymes is reported. The approach is based on the site-specific incorporation of a clickable non-natural amino acid into enzymes and two compatible click reactions. The multi-enzyme reaction system exhibited enhanced catalytic efficiency over the respective free enzymes.

No abstract

Rare earth elements (REEs) have become increasingly important materials owing to their use in the high-tech and clean-energy industries. However, the unpredictable supply, possible health risks, and environmentally unsustainable extraction practices associated with REEs have encouraged the development of green technologies for the selective extraction and recovery of metals. This study presents a simple and innovative approach for the selective extraction and recovery of total REEs. Elastin-like polypeptide (ELP) and the REE-binding domain (lanmodulin) are fused to form REEs-sensitive and thermo-responsive genetically encoded ELP called RELP, where ELP offered a reversible, inverse phase transition for repeated uses. The RELP are purified and used for the selective extraction of total REEs from competing non-REEs metals by controlling the solution temperature (4 and 37 °C) and pH. RELP exhibit high REE specificity, even in the presence of non-REE metal ions. The bound REEs are readily recovered during at least six repeated cycles, and the efficiency is maintained. Moreover, REEs are selectively recovered by RELP from steel slag leachate, a potential industrial source of REEs. RELP offers a rapid, selective, and scalable method for REE extraction and recovery. This technology can be adapted to recover other precious metals and commodities.

The number of therapeutic peptides for human treatment is growing rapidly. However, their development faces two major issues: the poor yield of large peptides from conventional solid-phase synthesis, and the intrinsically short serum half-life of peptides. To address these issues, we investigated a platform for the production of a recombinant therapeutic peptide with an extended serum half-life involving the site-specific conjugation of human serum albumin (HSA). HSA has an exceptionally long serum half-life and can be used to extend the serum half-lives of therapeutic proteins and peptides. We used glucagon-like-peptide 1 (GLP-1) as a model peptide in the present study. A “clickable” non-natural amino acid—p-azido-l-phenylalanine (AzF)—was incorporated into three specific sites (V16, Y19, and F28) of a GLP-1 variant, followed by conjugation with HSA through strain-promoted azide–alkyne cycloaddition. All three HSA-conjugated GLP-1 variants (GLP1_16HSA, GLP1_19HSA, and GLP1_28HSA) exhibited comparable serum half-lives in vivo. However, the three GLP1_HSA variants had different in vitro biological activities and in vivo glucose-lowering effects, demonstrating the importance of site-specific HSA conjugation. The platform described herein could be used to develop other therapeutic peptides with extended serum half-lives.