Protein hot spots at bio-nano interfaces

Chem. Commun., 2015

The combination of the very different chemical natures of carbon nanotubes (CNTs) and proteins gives rise to systems with unprecedented performance, thanks to a rich pool of very diverse chemical, electronic, catalytic and biological properties. Here we review recent advances in the field, including innovative and imaginative aspects from a nanoscale point of view. The tubular nature of CNTs allows for internal protein encapsulation, and also for their external coating by protein cages, affording bottom-up ordering of molecules in hierarchical structures. To achieve such complex systems it is imperative to master the intermolecular forces between CNTs and proteins, including geometry effects (e.g. CNT diameter and curvature) and how they translate into changes in the local environment (e.g. water entropy). The type of interaction between proteins and CNTs has important consequences for the preservation of their structure and, in turn, function. This key aspect cannot be neglected during the design of their conjugation, be it covalent, non-covalent, or based on a combination of both methods. The review concludes with a brief discussion of the very many applications intended for CNT-protein systems that go across various fields of science, from industrial biocatalysis to nanomedicine, from innovative materials to biotechnological tools in molecular biology research.

Functional integration of proteins with carbon-based nanomaterials holds great promise in emerging electronic and optoelectronic applications. Control over protein attachment poses a major challenge for consistent and useful device fabrication, especially when utilizing single/few molecule properties. Here, we exploit genetically encoded phenyl azide photochemistry to define the direct covalent attachment of three different proteins, including the fluorescent protein GFP, to carbon nanotube side walls. Single molecule fluorescence revealed that GFP attachment to SWCNTs changed fluorescence in terms of intensity and improved resistance to photobleaching; essentially GFP is fluorescent for much longer on attachment. The site of attachment proved important in terms of electronic impact on GFP function, with the attachment site furthest from the functional center having the larger effect on fluorescence. Our approach provides a versatile and general method for generating intimate protein-CNT hybrid bioconjugates. It can be potentially applied easily to any protein of choice; attachment position and thus interface characteristics with the CNT can easily be changed by simply placing the phenyl azide chemistry at different residues by gene mutagenesis. Thus, our approach will allow consistent construction and modulate functional coupling through changing the protein attachment position.

Accounts of Chemical Research, 2013

I ntegrating carbon nanotubes (CNTs) with biological systems to form hybrid functional assemblies is an innovative research area with great promise for medical, nanotechnology, and materials science applications. The specifics of molecular recognition and catalytic activity of proteins combined with the mechanical and electronic properties of CNTs provides opportunities for physicists, chemists, biologists, and materials scientists to understand and develop new nanomachines, sensors, or any of a number of other molecular assemblies. Researchers know relatively little about the structure, function, and spatial orientation of proteins noncovalently adsorbed on CNTs, yet because the interaction of CNTs with proteins depends strongly on the tridimensional structure of the proteins, many of these questions can be answered in simple terms. In this Account, we describe recent research investigating the properties of CNT/protein hybrids. Proteins act to solvate CNTs and may sort them according to diameter or chirality. In turn, CNTs can support and immobilize enzymes, creating functional materials. Additional applications include proteins that assemble ordered hierarchical objects containing CNTs, and CNTs that act as protein carriers for vaccines, for example. Protein/CNT hybrids can form bioscaffolds and can serve as therapeutic and imaging materials. Proteins can detect CNTs or coat them to make them biocompatible. One of the more challenging applications for protein/CNT hybrids is to make CNT substrates for cell growth and neural interfacing applications. The challenge arises from the structures' interactions with living cells, which poses questions surrounding the (nano)toxicology of CNTs and whether and how CNTs can detect biological processes or sense them as they occur. The surface chemistry of CNTs and proteins, including interactions such as πÀπ stacking interactions, hydrophobic interactions, surfactant-like interactions, and chargeÀπ interactions, governs the wealth of structures, processes, and functions that appear when such different types of molecules interact. Each residue stars in one of two main roles, and understanding which residues are best suited for which type of interaction can lead to the design of new hybrids. Nonlocally, the peptide or protein primary, secondary, and tertiary structures govern the binding of proteins by CNTs. The conjugation of proteins with CNTs presents some serious difficulties both experimentally and culturally (such as bridging the "jargon barrier" across disciplines). The intersection of these fields lies between communities characterized by distinctly different approaches and methodologies. However, the examples of this Account illustrate that when this barrier is overcome, the exploitation of hybrid CNTÀprotein systems offers great potential.

Nano Letters, 2004

Bacterial pili are nanofibers made of protein subunits. Here we report the controlled assembly of protein nanotubes from an engineered Pseudomonas aeruginosa type IV pilin monomer. The nanotubes are up to 100 µm long with an outer diameter of ∼6 nm and a predicted inner diameter of ∼2 nm. Protein nanotube formation appears to proceed through a hydrophobe-initiated conformational shift in the pilin monomer, which then self-associates to form thin linear filaments that coalesce to form long protein nanotubes. Protein nanotubes are highly attractive for nanotechnology, as protein engineering confers unprecedented control over mechanical and chemical properties. Moreover, like type IV pili, our nanotubes bind DNA, further broadening their appeal in nanotechnological applications.

Journal of Materials Research, 2015

Chemistry - A European Journal, 2012

Journal of Bionanoscience, 2007

The type IV pili of Pseudomonas aeruginosa are essentially protein nanofibres composed of multiple copies of a single pilin subunit. Type IV pili extend from the bacterial surface, and mediate specific adherence to biotic and abiotic surfaces. While deletion of the N-terminal region of the pilin's-helix allows for the ready expression of a highly soluble monomeric pilin protein, incubation of the monomeric protein with undecanethiol results in pilin oligomerization into protein nanotubes. In the present study, the ability of pilin-derived protein nanotubes to bind to grade 304 stainless steel surfaces was evaluated. Protein nanotubes bound to stainless steel with high affinity. Protein nanotube surface binding was observed to be a tip-associated event through competitive inhibition with a synthetic peptide corresponding to the pilin's C-terminal receptor binding domain. AFM studies established that the protein nanotubes utilize the pilin receptor binding domain to directly interact with the steel surface, demonstrating a 2-fold higher adhesive force for grain boundaries than for regions within grains. The adhesive force of the pilin receptor binding domain with the steel surface was determined by two methods and was conservatively estimated to be in the order of 26-55 pN/molecular interaction. Direct, specific binding of protein nanotubes, and/or receptor binding domain composite materials to a steel surface generates a novel metallo-biomolecular interface that forms preferentially on grain boundaries, enhancing the potential for these unique nanostructures in the development of biologically amenable nanosystems.

Scientific Reports, 2016

The study of proteins confined on a surface has attracted a great deal of attention due to its relevance in the development of bio-systems for laboratory and clinical settings. In this respect, organic bioelectronic platforms can be used as tools to achieve a deeper understanding of the processes involving protein interfaces. In this work, biotin-binding proteins have been integrated in two different organic thin-film transistor (TFT) configurations to separately address the changes occurring in the proteinligand complex morphology and dipole moment. This has been achieved by decoupling the output current change upon binding, taken as the transducing signal, into its component figures of merit. In particular, the threshold voltage is related to the protein dipole moment, while the field-effect mobility is associated with conformational changes occurring in the proteins of the layer when ligand binding occurs. Molecular Dynamics simulations on the whole avidin tetramer in presence and absence of ligands were carried out, to evaluate how the tight interactions with the ligand affect the protein dipole moment and the conformation of the loops surrounding the binding pocket. These simulations allow assembling a rather complete picture of the studied interaction processes and support the interpretation of the experimental results.

Current Opinion in Chemical Biology, 2010

Covalent and non-covalent conjugation of proteins to nanoparticles provides access to functional hybrid systems with applications in biotechnology, medicine, and catalysis. The creation of effective conjugates requires the retention of protein structure and function, a challenging task. In this review we discuss successes, challenges and opportunities in the area of protein-nanoparticle bioconjugation.

Acta biomaterialia, 2016

In the present work we investigate the key factors involved in the interaction of small-sized charged proteins with TiO2 nanostructures, i.e. albumin (negatively charged), histone (positively charged). We examine anodic nanotubes with specific morphology (simultaneous control over diameter and length, e.g. diameter - 15, 50 or 100 nm, length - 250 nm up to 10 μm) and nanopores. The nanostructures surface area has a direct influence on the amount of bound protein, nonetheless the protein physical properties as electric charge and size (in relation to nanotopography and biomaterial's electric charge) are crucial too. The highest quantity of adsorbed protein is registered for histone, for 100 nm diameter nanotubes (10 μm length) while higher values are registered for 15 nm diameter nanotubes when normalizing protein adsorption to nanostructures' surface unit area (evaluated from dye desorption measurements) - consistent with theoretical considerations. The proteins presence on ...