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The preferred spatial orientation of single-wall carbon nanotubes (SWCNTs) in their interaction with enzymes determines their behavior either as nano-supports or as inhibitors. alpha-chymotrypsin (alpha-CT) is considered a serine protease model for studying nanomaterial/proteases interactions. The interaction of alpha-CT with pristine single-wall carbon nanotubes is still unknown. Here alpha-CT/SWCNT hybrids are synthesized and characterized. Spectroscopic, microscopic and kinetic measurements, coupled to molecular dynamics simulations, provide a detailed description of the interaction between alpha-CT and SWCNTs. The SWCNT binding pocket was unambiguously identified. A perfect match is observed with the crevice structure of the alpha-CT substrate binding pocket. The activity of alpha-CT, upon SWCNT binding, is dramatically reduced, as expected by the interaction of the SWCNT in the active site of the protein. it-it stacking between aromatic residues and the conjugated surface of SWCNT governs alpha-CT/SWCNT interactions. An important role in the bonding appears also for purely hydrophobic residues and with residues able to establish surfactant-like interactions. The secondary structure of alpha-CT and the catalytic triad structure are not perturbed by the complex formation, on the contrary the volume of the substrate binding pocket is strongly reduced by SWCNT binding because SWCNT occupies the alpha-CT substrate binding site, clogging the active site. (C) 2020 Elsevier Inc. All rights reserved.

Carbon nanotubes, CNTs, and proteins could not be more chemically and physically distant. CNTs are hardly soluble and strongly hydrophobic. They are smooth wires, mechanically strong, with electronic properties that make them unique. Many proteins are water soluble and hydrophilic. They are highly corrugated and elastic. They can recognize a target molecule with high selectivity and sensitivity and can have catalytic activity. Integration of the remarkably different features of CNTs and proteins would create a new class of multifunctional materials. The conundrum to solve lies in how to best match proteins and CNTs. High-throughput virtual screening is used to predict the ability of 1207 proteins to recognize carbon tubes with a well-defined 1.3 nm diameter. The propensity for formation of protein-CNT hybrids is ranked. Experiments carried out in this work validated the computational results and show that the identified proteins are able to bind and disperse in water the selected CNTs. The highest scoring proteins are further examined in detail to identify general rules for binding and discussed for a variety of practical applications.

Photodynamic therapy (PDT) is considered a very promising therapeutic modality for antimicrobial therapy. Although several studies have demonstrated that Gram-positive bacteria are very sensitive to PDT, Gram-negative bacteria are more resistant to photodynamic action. This difference is due to a different cell wall structure. Gram-negative bacteria have an outer cell membrane containing lipopolysaccharides (LPS) that hinder the binding of photosensitizer molecules, protecting the bacterial cells from chemical attacks. Combination of the lipopolysaccharides-binding activity of Concanavalin A (ConA) with the photodynamic properties of Rose Bengal (RB) holds the potential of an innovative protein platform for targeted photodynamic therapy against Gram-negative bacteria. A ConA-RB bioconjugate was synthesized and characterized. Approximately 2.4 RB molecules were conjugated per ConA monomer. The conjugation of RB to ConA determines a decrease of the singlet oxygen generation and an increase of superoxide and peroxide production. The photokilling efficacy of the ConA-RB bioconjugate was demonstrated in a planktonic culture of E. coli. Irradiation with white light from a LED lamp produced a dose-dependent photokilling of bacteria. ConA-RB conjugates exhibited a consistent improvement over RB (up to 117-fold). The improved uptake of the photosensitizer explains the enhanced PDT effect accompanying increased membrane damages induced by the ConA-RB conjugate. The approach can be readily generalized (i) using different photo/sonosensitizers, (ii) to target other pathogens characterized by cell membranes containing lipopolysaccharides (LPS).