Direct and Selective Immobilization of Proteins by Means of an Inorganic Material-Binding Peptide: Discussion on Functionalization in the Elongation to Material-Binding Peptide
ABSTRACT Using an artificial peptide library, we have identified a peptide with affinity for ZnO materials that could be used to selectively accumulate ZnO particles on polypropylene-gold plates. In this study, we fused recombinant green fluorescent protein (GFP) with this ZnO-binding peptide (ZnOBP) and then selectively immobilized the fused protein on ZnO particles. We determined an appropriate condition for selective immobilization of recombinant GFP, and the ZnO-binding function of ZnOBP-fused GFP was examined by elongating the ZnOBP tag from a single amino acid to the intact sequence. The fusion of ZnOBP with GFP enabled specific adsorption of GFP on ZnO substrates in an appropriate solution, and thermodynamic studies showed a predominantly enthalpy-dependent electrostatic interaction between ZnOBP and the ZnO surface. The ZnOBP's binding affinity for the ZnO surface increased first in terms of material selectivity and then in terms of high affinity as the GFP-fused peptide was elongated from a single amino acid to intact ZnOBP. We concluded that the enthalpy-dependent interaction between ZnOBP and ZnO was influenced by the presence of not only charged amino acids but also their surrounding residues in the ZnOBP sequence.
- [Show abstract] [Hide abstract]
ABSTRACT: Adsorption behavior of a gold binding peptide was experimentally studied to obtain kinetics and thermodynamics parameters towards understanding of the binding of a peptide onto a solid metal surface. The gold-binding peptide, GBP1, was originally selected using a cell surface display library and contains 14-amino acid residues. In this work single- and three-repeats of GBP1 were used to assess the effects of two parameters: molecular architecture versus secondary structure on adsorption onto gold substrate. The adsorption measurements were carried out using surface plasmon resonance (SPR) spectroscopy at temperatures ranging from 10 ºC through 55 ºC. At all temperatures, two different regimes of peptide adsorption were observed which, based on the model, correspond to two sets of thermodynamics values. The values of enthalpy, Hads, and entropy, Sads, in these two regimes were determined using the Van't Hoff approach and Gibbs-Helmholtz relationship. In general, the values of enthalpy for both peptides are negative indicating GBP1 binding to gold is an exothermic phenomenon and that the binding of three repeat gold binding peptide (3l-GBP1) is about five times tighter than that for the single repeat (l-GBP1). More intriguing result is that the entropy of adsorption for the 3l-GBP1 is negative (-43.4±8.5 cal/mol K) while that for the l-GBP1 is positive (10.90±1.3 cal/mol K). Among a number of factors that synergistically contribute to the decrease of entropy, long-range ordered self-assembly of 3l-GBP1 on gold surface is the most effective, probably through both peptide-solid and peptide-peptide intermolecular interactions. Additional adsorption experiments were conducted in the presence of 2,2,2-trifluoroethanol (TFE) to determine how the conformational structures of the biomolecules responded to the environmental perturbation. We found that the peptides differ in their conformational responses to the change in solution conditions; while l-GBP does not fold in the presence of TFE, 3l-GBP1 adopted two types of secondary structure (-strand, -helix) and that peptide's binding to the solid is enhanced by the presence of low percentages of TFE solvent. These kinetics and thermodynamics results not only provide an example of adsorption behavior and binding of a genetically engineered peptide for an inorganic material (GEPI) but they also provide considerable insights into the fundamental understanding of the molecular recognition the peptides and their selective specificity for the solids. Moreover, comprehensive work described herein suggests that multiple repeat forms of the solid binding peptides possess a conformational component that can be exploited to further tailor affinity and binding of a given sequence to a solid material and ordered self-assembly, convenient tool in future practical applications.Biomacromolecules 06/2014; · 5.79 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Depositing gold nanoparticles is of great interest because of the many potential applications of nanoparticle films; however, generating dense surface nanoparticle coverage remains a difficult challenge. Using dual-affinity peptides we have synthesized gold nanoparticles and then pre-aggregated the particles in solution via interactions with metal ions. These nanoparticle aggregates were then deposited onto silicon dioxide surfaces using another dual-affinity peptide to control binding to the substrate. The results demonstrate that when divalent ions like Zn2+ or Ni2+ are used, densely packed gold nanoparticle monolayers are formed on the silicon dioxide substrate, which may have applications in fields like molecular electronics.Applied Surface Science 03/2014; 296:24–30. · 2.54 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: The growing usage of nanoparticles of zinc sulfide as quantum dots and biosensors calls for a theoretical assessment of interactions of ZnS with biomolecules. We employ the molecular-dynamics-based umbrella sampling method to determine potentials of mean force for 20 single amino acids near the ZnS (110) surface in aqueous solutions. We find that five amino acids do not bind at all and the binding energy of the remaining amino acids does not exceed 4.3 kJ/mol. Such energies are comparable to those found for ZnO (and to hydrogen bonds in proteins) but the nature of the specificity is different. Cysteine can bind with ZnS in a covalent way, e.g., by forming the disulfide bond with S in the solid. If this effect is included within a model incorporating the Morse potential, then the potential well becomes much deeper-the binding energy is close to 98 kJ/mol. We then consider tryptophan cage, a protein of 20 residues, and characterize its events of adsorption to ZnS. We demonstrate the relevance of interactions between the amino acids in the selection of optimal adsorbed conformations and recognize the key role of cysteine in generation of lasting adsorption. We show that ZnS is more hydrophobic than ZnO and that the density profile of water is quite different than that forming near ZnO-it has only a minor articulation into layers. Furthermore, the first layer of water is disordered and mobile.The Journal of Chemical Physics 03/2014; 140(9):095101. · 3.12 Impact Factor