Enzyme catalysis within cells not only enables in-situ monitoring of cellular metabolism for disease diagnosis, but also offers promising therapeutic benefits for defective enzyme-induced diseases. Hydrogen-bonded organic frameworks (HOFs) is well suited to serve as a general platform for encapsulation of enzymes to facilitate intracellular biocatalysis.

In a study recently published in Angewandte Chemie International Edition, the research group led by Prof. WANG Ming from Institute of Chemistry of the Chinese Academy of Sciences developed a universal strategy for enzyme incorporation into nanoscale HOFs through in-situ self-assembly of proteins and HOFs precursors, which further showed preserved enzymatic activity inside living cells.

The researchers firstly prepared nanosized HOFs via self-assembly of tetrakis(4-amidiniumphenyl)methane (T) and azobenzenedicarboxylate (A) in aqueous solution at room temperature termed as TA-HOFs. By optimizing the concentration of HOFs motifs and proteins, they then obtained nanoscale protein-encapsulated biocomposites which showed preserved protein structure and function. The as-prepared protein@TA-HOFs can deliver protein into cells without scarfing protein function.

Using green fluorescent protein (GFP) as a proof of concept, the researchers found that GFP@TA-HOFs showed high uptake efficiency by human cervical carcinoma (HeLa) cells with minor cellular toxicity, and confirmed efficient endo/lysosome escape by counterstaining of endosome. Notably, the in-situ self-assembly strategy was generally applicable to proteins with different surface charges and sizes to prepare nanoscale pretein@biocomposites. TA-HOFs can efficiently deliver highly surface charged GFPs, proving the universal of the TA-HOFs cellular delivery platform.

Besides, the researchers studied intracellular delivery of enzymes for enzymatic catalysis inside cells. They found that TA-HOFs can deliver enzymes such as horseradish peroxidase (HRP) and β-galactosidase into HeLa cells with preserved enzyme activity, and intracellular delivery of catalase (CAT) into neural cells (SH-SY5Y) can alleviate reactive oxygen species (ROS) accumulation inside cells, which is one of the causes of most neurodegenerative diseases.

This study paved the way for applying HOFs as a novel universal platform for biomacromolecule delivery and disease treatment, and was highlighted in Nature Reviews Chemistry.