Mechanism Of Protein Adsorption Research Articles

Mechanisms of human plasma protein adsorption on the surface of perfluorocarbon emulsion stabilized with proxanol 268

It has been shown that sorption of most proteins with the molecular weight lower than 200 kDa from human blood plasma on the surface of perfluorocarbon emulsion, stabilized with proxanol 268, is mainly based on hydrophobic interaction, whereas sorption of immunoglobulin G is mainly the result of electrostatic interaction. The removal of lipidic components from plasma leads to the increase of a total amount of adsorbed proteins by 35%. Particularly, when lipidic components are removed, sorption of apolipoprotein AI and immunoglobulin G is considerably bettered as well as sorption of other proteins with the molecular weight of about 50 and 60 kDa occurs. It has been out that apolipoprotein AI in the adsorbed condition loses its capability of tryptophan fluorescence, which might be probably determined by the quenching influence of the perfluorocarbon core of nanoparticle. We think that the findings obtained also indicates considerable conformational rearrangements of this protein during adsorption. It was shown, that the fluorescence of proteins with sorption on nanoparticles in emulsion based on the hydrophobic interaction, is completely or partially quenched.

Protein adsorption kinetics from single- and binary-solution

Comparison of protein mass-adsorption-rates to rates-of-change in interfacial tensions reveals that mass adsorption is decoupled from interfacial energetics. This implies that energy-barrier theories describing protein-adsorption kinetics do not accurately capture the physics of the process. An alternative paradigm in which protein molecules rapidly diffuse into an inflating interphase which subsequently slowly shrinks in volume, concentrating adsorbed protein and causing slow concomitant decrease in interfacial tensions, is shown to be consistent with adsorption kinetics measured by solution depletion and tensiometry. Mass adsorption kinetics observed from binary-protein solution is compared to adsorption kinetics from single-protein solution, revealing that organization of two different-sized proteins within the interphase can require significantly longer than that adsorbed from single-protein solution and may require expulsion of initially adsorbed protein which is not observed in the single-protein case.

General Description of the Adsorption of Proteins at Their Iso-electric Point in Nanoporous Materials

A simple but remarkably precise geometric pore-filling model is proposed and experimentally validated for the adsorption of proteins at their iso-electric point (pI) in nanoporous materials. Three different globular proteins-lysozyme, myoglobin, and bovine serum albumin-are used as model proteins to study protein adsorption on two types of ordered mesoporous materials-silica and carbon-which allows us to study the effects of protein and surface structure on the protein adsorption mechanism. The geometric pore-filling model confirms that proteins are closely packed inside the pore channels of mesoporous materials, leading to an exceptionally large protein loading capacity. A relationship for the amount of adsorbed protein as a function of protein size, nanopore volume, and pore diameter is derived. The pore space gradually fills up to complete packing of the available pore space at the highest protein concentration. The high precision of the geometric pore-filling model demonstrates its utility to predict the protein adsorption capacity of ordered nanoporous materials.

Energetics of protein adsorption on amine-functionalized mesostructured cellular foam silica

The energetics of lysozyme adsorption on aminopropyl-grafted MCF silica (MCF-NH2) are compared to the trends observed during lysozyme adsorption on native MCF silica using flow microcalorimetry (FMC). Surface modification on MCF silica affects adsorption energetics significantly. All thermograms consist of two initial exothermic peaks and one later endothermic peak, but the heat signal trends of MCF-NH2 are opposite from those observed for adsorption onto native MCF silica in salt solutions of sodium acetate and sodium sulfate. At low ionic strength (0.01 M), LYS adsorption onto MCF-NH2 was accompanied by a large exotherm followed by a desorption endotherm. With increasing ionic strength (0.1 and 3.01 M), the magnitude of the thermal signal decreased and the total process became less exothermic. Also a higher protein loading of 14 μmol g −1 was obtained at low ionic strength in batch adsorption isotherm measurements. Taken together, the FMC thermograms and batch adsorption isotherms reveal that MCF-NH2 has the nature of an ion exchange adsorbent, even though lysozyme and the aminopropyl ligands have like net charges at the adsorption pH. Reduced electrostatic interaction, reduced Debye length, and increased adsorption-site competition attenuate exothermicity at higher ionic strengths. Thermograms from flow microcalorimetry (FMC) give rich insight into the mechanisms of protein adsorption. A two-step adsorption mechanism is proposed in which negatively charged surface amino acid side chains on the lysozyme surface make an initial attachment to surface aminopropyl ligands by electrostatic interaction (low ionic strength) or van der Waals interaction (high ionic strength). Secondary attachments take place between protruding amino acid side chains and silanol groups on the silica surface. The reduced secondary adsorption heat is attributed to the inhibitory effect of the enhanced steric barrier of aminopropyl group on MCF silica.

Microcalorimetric study of the adsorption of PEGylated lysozyme on a strong cation exchange resin

Abstract Adsorption of native as well as mono-, di-, and tri-PEGylated lysozyme on Toyopearl Gigacap S-650M in sodium phosphate buffer is studied by isothermal titration calorimetry and by independent adsorption equilibrium measurements at pH 6 and 25 °C. The production and separation of PEGylated lysozyme is discussed. Two different PEG sizes are used (5 kDa and 10 kDa) which leads to six different forms of PEGylated lysozyme which were systematically studied. The sodium chloride concentration is varied according to the elution conditions in the production process. The specific enthalpy of adsorption Δ h p ads is determined from the calorimetric and the adsorption equilibrium data. It was found to be exothermal and constant with increasing adsorber loading for native lysozyme. For all PEGylated forms there is an influence of the adsorber loading on Δ h p ads which is found to become more important with increasing PEGylation degree (total molecular weight of conjugated PEG). At low loadings the adsorption of all PEGylated lysozyme forms is exothermal. With increasing loading the adsorption becomes less exothermal and for the species with higher PEGylation degree also endothermal effects are observed at higher loadings. A thermodynamic analysis is carried out by which the enthalpic and entropic contributions to the binding constants are quantified. The findings are discussed on a molecular level. The results provide insight into the adsorption mechanisms of polymer-modified proteins on chromatographic resins.

Interactions of nanoparticles with plasma proteins: implication on clearance and toxicity of drug delivery systems

Introduction: Intravenously injected nanoparticles, like any other foreign pathogen that enters the body, encounter multiple lines of defense intended to neutralize and eliminate the invading substance. Adsorption of plasma proteins on the nanoparticle surface is the first barrier of defense, which could lead to physical changes in the formulation, such as aggregation and charge neutralization, biochemical activation of defense cascades, and trigger elimination by multiple types of phagocytic cell.Areas covered: In this review, recent knowledge on the mechanisms that govern the interactions of nanoparticles (micelles, liposomes, polymeric and inorganic nanoparticles) with plasma proteins is discussed. In particular, the role of the nanoparticle surface properties and protective polymer coating in these interactions is described. The mechanisms of protein adsorption on different nanoparticles are analyzed and the implications on the clearance, toxicity and efficacy of drug delivery are discussed. The review provides readers with the biological insight into the plasma/blood interactions of nanoparticles.Expert opinion: The immune recognition of nanoparticles can seriously affect the drug delivery efficacy and toxicity. There is at present not enough knowledge on the mechanisms that dictate the nanoparticle immune recognition and stability in the biological milieu. Understanding the mechanisms of recognition will become an important part of nanoparticle design.

Single‐molecule imaging of protein adsorption mechanisms to surfaces

Protein-surface interactions cause the desirable effect of controlled protein adsorption onto biodevices as well as the undesirable effect of protein fouling. The key to controlling protein-surface adsorptions is to identify and quantify the main adsorption mechanisms: adsorptions that occur (1) while depositing a protein solution onto dry surfaces and (2) after the deposition onto wet surfaces. Bulk measurements cannot reveal the dynamic protein adsorption pathways and thus cannot differentiate between the two adsorption mechanisms. We imaged the interactions of single streptavidin molecules with hydrophobic fused-silica surfaces in real-time. We observed both adsorbed proteins on surfaces and diffusing proteins near surfaces and analyzed their adsorption kinetics. Our analysis shows that the protein solution deposition process is the primary mechanism of streptavidin adsorption onto surfaces at the subnanomolar to nanomolar protein concentrations. Furthermore, we found that hydrophilic fused-silica surfaces can prevent the adsorption of streptavidin molecules.

Mixed-SAM Surfaces Monitoring CTX-Protein Part I: Using Atomic Force Microscope Measurements

Fast and efficient detection of Cobra cardiotoxin (CTX) protein molecules on biochip surfaces is an example of application in biotechnology. One potential application of mixed self assembled monolayers (SAMs) as chip surfaces yield different binding affinities of the CTX proteins, a series of studies on the interaction force between CTX proteins and the mixed SAMs surfaces formed from mixtures of two thiols with the same/different chain lengths and/or with the same/different terminal groups will be investigated. In these dual papers, the mixed SAMs of n-alkinethiol SAMs of different chain lengths are chosen as the first examples of this series due to the simple functions of the mixed SAMs surface structure. Thus, the adhesion force measurements of CTX protein molecules on mixed SAMs of n-alkinethiol SAMs of different chain lengths: 1-decanethiol (C9) and 1-hexanethiol (C5) with different mixing ratios are developed and conducted using atomic force microscope (AFM). There are two major tasks in Part I of the dual papers: the development of the AFM measurements providing reliable information, and selection of the surface with highest binding affinity among this mixed SAMs group. Results indicate that the adhesion forces for CTX protein molecules on mixed SAMs with mixing ratio (χ(C9)) of 0.25, 0.5, 0.75 and 1, are 1.26, 1.8, 1.38, and 1.25 folds respectively, compared with the adhesion force of CTX protein molecules on the C5 surface only. Therefore, the SAM surfaces of χ(C9) = 0.5 is the best choice as a biomaterial sensor of this group of mixed SAMs because the strongest binding force and highest efficiency. Effects of the loading force of the AFM operation, the radius of curvature of the AFM tip, and the AFM tip endurance as well as control experiments were examined to ensure the quantitative determination of adhesion force for AFM measurement. The physical mechanism of protein adsorption on SAM surfaces will be studied and analyzed by molecular dynamics (MD) simulations and will be reported in Part II of the dual papers to compensate the limited information on the interaction taking place at atomic level that experiments cannot provide.

On the Mechanism of Protein Adsorption onto Hydroxylated and Nonhydroxylated TiO2 Surfaces

Protein adsorption onto implant surfaces is of great importance for the regulation of implant bioactivity. Surface modification of implants is a promising way in the molecular design of biocompatib ...

Thermal Oxidation for Controlling Protein Interactions with Porous Silicon

Thermal oxidation of porous silicon (pSi) has been used to control interactions with three proteins; lysozyme, papain, and human serum albumin (HSA) enabling the influences of protein structure, molecular weight, and charge to be elucidated. Adsorption behavior was assessed via adsorption isotherms while the structures of adsorbed proteins were investigated using a bioactivity assay, FTIR, and zeta potential. Time-of-flight secondary ion mass spectrometry was used to examine protein pore penetration. High protein adsorption onto unoxidized pSi (240-610 microg/m(2)) was attributed to predominately hydrophobic interactions which resulted in structural changes of the adsorbed proteins and significant loss of bioactivity. Thermal oxidation at 400 and 800 degrees C significantly reduced protein adsorption (80-485 microg/m(2)) by reducing hydrophobicity. Oxidation of pSi modified the protein adsorption mechanisms to solely electrostatic attraction for positively charged proteins and structural rearrangement for negatively charged proteins. Adsorption via electrostatic attraction preserved protein bioactivity and zeta potential, thus inferring a retention of their native structure. In contrast, the negative charge and globular structure of HSA resulted in a loss of structure. We have demonstrated that thermal oxidation of pSi can be used to control protein interactions, adsorbed structure, and bioactivity.

Study of the adsorption process of bovine serum albumin on passivated surfaces of CoCrMo biomedical alloy

Adsorption of bovine serum albumin (BSA) on CoCrMo surface was studied by electrochemical techniques in order to determine the mechanism of protein adsorption at different surface conditions (which are reached by considering different passivation times) in phosphate buffered solution (PBS). At open circuit potential (OCP), adsorption kinetic was influenced by surface passivation when passivation time was lower than 1 h, whereas, at higher passivation times, no apparent modification in the kinetic mechanisms of adsorption was observed. On the other hand, at a passive applied potential, the BSA addition decreased the passivation kinetics of the process at all the passivation times.

Influence of Surface Conditions on the Adsorption Process of Bovine Serum Albumin (BSA) on CoCrMo Biomedical Alloy

CoCrMo alloys are biocompatible materials widely used for orthopaedic implants. The biocompatibility of CoCrMo alloy is closely related to its excellent corrosion resistance, imparted by a passive oxide film that forms spontaneously on the alloy surface. The adsorption of organic species present in the synovial fluids may attach with this passive film and modify their surface reactions. BSA adsorption on CoCrMo surface was studied in order to determine the mechanism of protein adsorption at different surface conditions and passivation times in phosphate buffered solution (PBS). At Open Circuit Potential (OCP), adsorption kinetics were influenced by surface passivation when passivation time was lower than 1 hour, whereas, at higher passivation times, no apparent modification in the kinetic mechanisms of adsorption was observed. On the other hand, at a passive applied potential, the BSA addition decreased the passivation kinetics of the process at all the passivation times.

Comparison of Surface Tension Components and Hansen Solubility Parameters Theories. Part I: Explanation of Protein Adsorption on Polymers

Surface tension components and Hansen solubility parameters are two different theories used to explain and predict the compatibility and miscibility for polymer blends, composite polymer materials, and additives in polymer matrix. In this paper, the adsorption mechanism of bovine serum albumin (BSA) on eight polymers (PC, POM, PBT, PPO, PSF, PVDF, PEI, and PES) was researched through surface tension components and Hansen solubility parameters theories. The adsorption amounts of BSA on the polymers were measured through static adsorption experiments. It was found that the BSA adsorption amount cannot be directly correlated with the Hansen solubility parameters. However, there is high correlation between the Lifshitz-van der Waals component of surface tension of the polymers and BSA adsorption amount. With the increase in van der Waals value, the BSA adsorption amount on the polymers increases, but there still exists a complicated behavior. The protein adsorption mechanism is complex.

Investigation of the mechanism of protein adsorption on ordered mesoporous silica using flow microcalorimetry

The adsorption of bovine serum albumin (BSA) and lysozyme (LYS) on siliceous SBA-15 with 24 nm pores was studied using flow microcalorimetry; this is the first attempt to understand the thermodynamics of protein adsorption on SBA-15 using flow microcalorimetry. The adsorption mechanism is a strong function of protein structure. Exothermic events were observed when protein–surface interactions were attractive. Entropy-driven endothermic events were also observed in some cases, resulting from lateral protein–protein interactions and conformational changes in the adsorbed protein. The magnitudes of the enthalpies of adsorption for primary protein–surface interactions decrease with increased surface coverage, indicating the possibility of increased repulsion between adsorbed protein molecules. Secondary exothermic events were observed for BSA adsorption, presumably due to secondary adsorption made possible by conformational changes in the soft BSA protein. These secondary adsorption events were not observed for lysozyme, which is structurally robust. The results of this study emphasize the influence of solution conditions and protein structure on conformational changes of the adsorbed protein and the value of calorimetry in understanding protein–surface interactions.

Co-adsorption of electrolyte and protein to Ag colloid observed by surface-enhanced Raman scattering

Protein detection using surface-enhanced Raman scattering (SERS) usually requires electrolytes to yield an enhanced SERS signal. However, the adsorption mechanism of electrolyte and protein to Ag colloid is not yet clearly understood. In this work, we have investigated co-adsorption of NO(3)(-) and lysozyme to Ag colloid using SERS. Three experimental factors including concentration of lysozyme (10(-5) and 10(-6) M), concentration of NO(3)(-) (0, 1, 2, 3 and 5 mM) and drying temperature (25 and 100 degrees C) have been studied. The results have shown that the co-adsorption of the adsorbates (lysozyme and NO(3)(-)) on a SERS substrate and the non-absorption of NO(3)(-) on the substrate can be controlled by using different experimental conditions. The co-adsorption manner of lysozyme and NO(3)(-) is consistent with the mechanism of double-layer adsorbates when a protein adsorbs on a solid/liquid interface. The variation in protein conformation, especially the main-chain conformation, seems to affect the adsorption manner of the adsorbates. It has been found that the final adsorption result is not affected by the addition sequence of lysozyme and NO(3)(-) during the sample preparation.

Cover Picture: Electrophoresis 12'09

AbstractIssue no. 12 is a special issue on “Bioanalysis” consisting of 2 reviews articles and 20 research articles. The two review articles deal with fundamental aspects and methodologies. The first review article addresses molecular mechanisms of protein adsorption on silica surfaces while the second review article is a survey of microdialysis techniques used in combination with subsequent separation and detection methods. The 20 research articles are distributed over 4 parts: (i) Nucleic Acids, (ii) Protein Analysis using CE and Microfluidics, (iii) Gel‐Based Protein Analysis and Proteomics Profiling, and (iv) Metabolic and Toxicological Profiling.The first paper in the issue is on “DNA migration mechanism analyses for applications in capillary and microchip electrophoresis”, and is the contribution from Annelise Barron to the Supplement Issue “Electrophoresis: Past, Present and Future”.The second paper in this issue is a Fast Track article on “Nanostructured Pillars Consist of Vertically Aligned Carbon Nanotubes as a Stationary Phase in Microcapillary Electrochromatography (?CEC)”.

Protein Adsorption on Hydroxyapatite Nanosensors with Different Crystal Sizes Studied In Situ by a Quartz Crystal Microbalance with the Dissipation Method

Hydroxyapatite (HAp) nanocrystals with different crystal sizes were deposited by the electrophoretic deposition method on the gold surface of a quartz crystal microbalance with a dissipation probe. The nanosensors formed this way were used to elucidate the adsorption mechanism of proteins with a similar pI value. The crystal sizes and the area of the a‐plane affected only the adsorption amount of human serum albumin, but not that of bovine plasma fibrinogen. The viscoelastic property, ΔD/Δf, of each absorbed layer on the nanosensors was almost constant. The protein adsorption mechanism can be explained as follows: the dissociated carboxyl groups (negative charge) of albumin were interacted with calcium ions and the hydrated amine groups (positive charge) at the αC domain of fibrinogen were with phosphate ions on the HAp surface.

Protein adsorption mechanisms on solid surfaces: lysozyme-on-mica

A methodology for discovering the mechanisms and dynamics of protein clustering on solid surfaces is reviewed and complemented by atomistic molecular dynamics (MD) simulations. In situ atomic force microscopy images of the early stages of protein film formation are quantitatively compared with Monte Carlo simulations, using cluster statistics to differentiate various growth models. We have studied lysozyme adsorption on mica as a model system, finding that all surface-supported clusters are mobile with diffusion constant inversely related to cluster size. Furthermore, our results suggest that protein monomers diffusing to the surface from solution only adhere to the bare surface with a finite probability. Fully atomistic MD simulations reveal that the lysozyme does indeed have a preferred orientation for binding to the surface, so that proteins with incorrect orientations move away from the surface rather than towards it. Agreement with experimental studies in the literature for the residues involved in the surface adsorption is found.

PH-induced changes in adsorbed β-lactoglobulin molecules measured using atomic force microscopy

We have used atomic force microscopy (AFM) imaging and force spectroscopy to study β-lactoglobulin (β-LG) food protein molecules adsorbed onto a mica surface. In particular, we have studied the effect of in situ changes in pH on several different properties: the topographical morphology of the adsorbed β-LG molecules, adhesion of the β-LG molecules to the underlying mica substrate, and the mechanical unfolding of the β-LG molecules. In AFM images, the structure of the adsorbed protein layer was observed to change dramatically with changes in pH. This result was consistent with the mechanical unfolding of single protein molecules within the adsorbed protein layer at different pH values performed using AFM. The short rupture length (∼50 nm) measured for the fully unfolded protein at an acidic pH value of 2.5 is in good agreement with the dominant single molecule population measured previously for this pH value. Unfolding β-LG molecules from the same protein layer at a neutral value of pH = 6.8 resulted primarily in longer rupture lengths, corresponding to dimers of β-LG. AFM force–distance curves collected at pH = 9 were dominated by a large repulsion between the AFM tip and the adsorbed protein layer, which is likely due to the extended nature of the molecules because of irreversible denaturation for pH values greater than 9. This work provides a novel insight into the mechanisms of protein adsorption onto surfaces and shows that AFM force spectroscopy is a promising tool for probing in situ conformational changes in single molecules under various conditions.

Protein interaction with hydrogel contact lenses.

The first event observed at the interface between a contact lens and tear fluid is protein adsorption. Tears have a rich and complex composition, allowing a wide range of interactions and competitive processes. The majority of studies avail-able on protein adsorption have focused on blood/plasma/serum protein on solid surfaces. However, recently the interaction of tear proteins with contact lenses has become an important field of research following the widespread use of contact lenses in many physiological and pathological conditions. Protein adsorption on contact lenses is the overall result of various types of in-teractions between the different components present, i.e. the chemical composition and the surface charge, the structure of the protein molecules, the nature of the medium (tears) and many other solutes present in tears. The chemical structure of contact lenses and their physical and biological behavior are considered, followed by tear film structure and biochemistry. Finally, the nature and mechanism of protein adsorption on contact lenses, factors affecting adsorption and attempts to reduce protein ad-sorption are reviewed. (Journal of Applied Biomaterials & Biomechanics 2004; 2: 1-19).