Binding Of BSA Research Articles

Interactions and characterization of Zein-BSA nanoparticles: Multi-spectral analysis and molecular simulations

Zein-bovine serum albumin (BSA) nanoparticles (NPs) with varying mass ratios were prepared via anti-solvent precipitation method at pH 4.0. Zein-BSA NPs at high Zein mass ratios exhibited superior particle characteristics and surface morphology. Ultraviolet-visible, fluorescence, and X-ray diffraction analyses demonstrated the formation and conformational transition during the binding of Zein and BSA, while ANS hydrophobic binding, fourier transform infrared spectra, and circular dichroism confirmed the hydrophobic exposure and secondary structure assignment. Molecular dynamic simulations revealed that Zein spontaneously bound to the IIB domain of BSA with a binding free energy of −63.52 ± 6.37 kcal/mol, which was dominated by hydrophobic and electrostatic interactions, supplemented by hydrogen bonds. Dissociation experiments supported the key role of hydrophobic and electrostatic interactions in maintaining the stability of the NPs. These findings provided insights into the formation and stabilization mechanisms of Zein-BSA NPs, and their potential applications as encapsulation carriers for bioactive substances or as novel nanoscale food substrates.

Binding of serum albumin to perfluorooctanoic acid reduced cytotoxicity

With the ubiquitous applications of perfluorinated compounds such as perfluorooctanoic acid (PFOA) in industrial and commercial products, the toxicity of these engineered materials in environmental and public health is received growing attention. As a typical organic pollutant, PFOA has been extensively found in wildlife and human bodies, and can preferentially bind to serum albumin in vivo. However, the importance of protein–PFOA interactions on the cytotoxicity of PFOA could not be stressed enough. In this study, we used both experimental and theoretical approaches, to investigate the interactions of PFOA with bovine serum albumin (BSA, the most abundant protein in blood). It was found that PFOA could mainly interact with Sudlow site I of BSA to form BSA-PFOA complex, in which van der Waals forces and hydrogen bonds played dominant roles. Moreover, the strong binding of BSA could greatly alter the cellular uptake and distribution of PFOA in human endothelial cells, and result in the decreases of reactive oxygen species formation and cytotoxicity for these BSA-coated PFOA. Consistently, the addition of fetal bovine serum into cell culture medium also significantly mitigated PFOA-induced cytotoxicity, which was attributed to the extracellular complexation between PFOA and serum proteins. Altogether, our study demonstrates that the binding of serum albumin to PFOA could reduce its toxicity by affecting the cellular responses.

Evaluation the binding of chlorogenic acid with bovine serum albumin: Spectroscopic methods, electrochemical and molecular docking

Chlorogenic acid(CGA) is the common active phenolic acid in Chinese medicinal materials such as honeysuckle and eucommia. It is a class of small molecules with multiple activities such as antioxidant, inhibiting cancer cells, lowering blood sugar and lowering blood pressure. In this paper, UV–vis spectroscopy, fluorescence spectroscopy, circular dichroism, molecular dynamics simulation and cyclic voltammetry (CV) electrochemical analysis were used to investigate the mechanism about interaction between CGA and BSA. Based on fluorescence quenching analysis, CGA quenched the inherent fluorescence of BSA remarkably through a static mechanism. The obtained value of binding constant (Kb = 5.75 × 105 L·mol−1) revealed a high binding affinity between CGA and BSA. The simulated molecular docking showed that hydrophobic force were also involved in the interaction between BSA and CGA. This paper also investigate the effect of temperature and metal ions on the binding of CGA and BSA. When the temperature increased, the binding of BSA and CGA was destroyed. Metal ions affect both the structure of BSA and the combination of BSA and CGA. By studying the mechanism of CGA interaction with BSA, we elucidated the storage and transport mechanism of CGA in vivo under simulated human environment and temperature conditions.

Constructing novel nanofibrous polyacrylonitrile (PAN)-based anion exchange membrane adsorber for protein separation

Purification of biopharmaceutical streams is essential for producing high quality therapeutic bioproducts. This work developed a novel polyacrylonitrile (PAN)-based nanofibrous membrane with strong anion exchange functionality via electrospinning. The key material functionality was obtained via RAFT copolymerization of acrylonitrile and dimethylaminoethyl acrylate (pAD), followed by quaternization to form quaternary amine (QA) ligands, namely pAQ, a series of nanofibrous PAN-pAQ membranes were electrospun by blending the pAQ copolymer with PAN homopolymer at varying ratios. The chemistry of the respective pAQ copolymer and resulting membranes was confirmed by NMR and FTIR, evidencing successful functionalization. As compared to the reference pure membrane PAN4 that was negatively charged, the resulting composite membranes showed a positive surface charge. The investigation on surface morphology revealed that the nanofiber diameter increased from 300 nm to 1 μm with an increasing blend ratio from 1:4 to 1:7 for the PAN-pAQ membranes. Such trend in surface micro/nano morphology changes strongly influenced other surface properties such as increased pore size, reduced specific surface area and increased hydrophobicity. The static binding of model protein BSA of PAN-pAQ membranes firstly increased with blend ratio from 1:4 to 1:5, and then decreased at 1:7, which was attributed to the complex trade-off relationship between surface micro/nano-structure and hence distribution/density of quaternary functional groups. The PAN-pAQ membranes showed about a 10-fold increase in static binding capacity compared to PAN4, up to 310–320 mg·g−1 at a blend ratio of 1:5. Thus through this study, we were able to demonstrate a facile route to incorporate pre-functionalized copolymers into conventional polymers to form chromatographic membranes, with many possibilities to tailor membrane functionality for a wide range of applications.

Thermodynamic analysis of albumin interaction with monosodium glutamate food additive: Insights from multi-spectroscopic and molecular docking approaches

Monosodium glutamate (MSG) is an example of food additive, which is used as a flavor enhancer in various foodstuffs. Because of its extensive use in food production, the toxicology and possible negative effects of MSG additive on albumin must be well investigated. Therefore, for the first time, the binding of BSA and MSG have been studied using multispectroscopic and molecular modeling approaches under physiological conditions. BSA fluorescence intensity has been diminished upon the addition of increased MSG concentrations. The Stern–Volmer quenching constant (KSV) value increase with rising temperature and bimolecular quenching rate constant of protein (kq) value was larger than 2.0 × 1010 L mol− 1 s− 1, which shows that the fluorescence quenching was both dynamic and static due to the BSA-MSG complex formation. The positive values for both ΔS0 (888.291 J mol−1K−1) and ΔH0 (243.903 kJ mol−1) proposed that the hydrophobic forces played predominant roles in the binding of MSG to BSA. Also, the negative value of ΔG0 indicated that the interaction between two compounds is spontaneous and enthalpy driven. The UV–VIS spectra results confirmed the binding of MSG to BSA. The circular dichroism analysis exhibited that BSA conformation change occur upon interaction with MSG. Moreover, molecular modeling analysis indicated that glutamate remarkably tends to bind into a cavity above Sudlow II with lowest calculated binding energy of - 8.24 kcal/mol.

Kinetic and thermodynamic insights into interaction of albumin with piperacillin: Spectroscopic and molecular modeling approaches

The current study investigated the binding of BSA and piperacillin at multifarious concentrations (1–128 μM) along with four different temperatures. To do this, three sensitive methods were employed including surface plasmon resonance (SPR), fluorescence spectroscopy and molecular docking (MD). In the SPR method, BSA immobilized on the carboxymethyl dextran (CMD) hydrogel sensor chip through NHS/EDC activation. The Van’t Hoff equation was applied for thermodynamic parameters determination at four distinct temperatures. Fluorescence spectroscopy results showed that binding constants decrease upon BSA interaction with piperacillin by rising temperature, which is indicative of BSA-piperacillin’s complex formation and fluorescence quenching mechanism of BSA induced by piperacillin is via both static and dynamic quenching process. The SPR analysis showed dose-response sensorgrams of BSA as piperacillin concentration increased. In this regard, piperacillin demonstrated a higher affinity to BSA that was concluded via a smaller amount of equilibrium constants (KD). Besides, docking results indicated that IB and IIIA subdomains of BSA are the most favorable sites for piperacillin binding. Therefore, the attained results showed that piperacillin interaction with BSA could be changed from van der Waals interactions to hydrogen bonds depending on the BSA subdomains.

Binding of l-tryptophan and bovine serum albumin by novel gold nanoparticles capped with amphiphilic sulfonatomethylated calixresorcinarenes

A simple, one-step synthesis of spherical gold nanoparticles (dcore=38 and 25nm) in aqueous solution has been carried out using sulfonatomethylated calixresorcinarenes with methyl (C1S) and pentyl (C5S) substituents on the lower rim both as reducing and stabilizing agents. The nanoparticles obtained, Au@C1S and Au@C5S, were characterized by absorption spectrophotometry, TEM, DLS, IR and TGA. The gold nanoparticles are stabilized by the interaction that occurs between macrocycles' sulfo-groups and the metal surface. It has been shown, that Au@C5S nanoparticles have a higher thermal stability than Au@C1S nanoparticles due to the denser packing of the macrocycle on the surface of gold nanoparticles caused by amphiphilic nature of the C5S macrocycle. C5S or Au@C5S associate cooperatively with l-tryptophan to form a nanoscale system capable of a spectral response to changing pH in aqueous solution.A detailed study of the interaction of С5S and Au@C5S with BSA in aqueous solution was carried out by spectrophotometry, DLS and fluorimetry. It was found that C5S and Au@C5S both form stable complexes with the protein, which was confirmed by the static character of the quenching of BSA fluorescence. The mechanism of the interaction of BSA with the macrocycles in aqueous solution and on the surface of gold nanoparticles was investigated. The quenching (KSV) and binding (Kas) constants, the number of binding sites (n), and thermodynamic interaction parameters (ΔG, ΔH, ΔS) were calculated. It was found that the process of protein binding by the free macrocycle and the modified gold nanoparticles is exothermic and spontaneous, and hydrogen bonding and van der Waals interactions make a major contribution into the binding. The synchronous fluorescence method showed that the binding of BSA with macrocycle influences the microenvironment of the tryptophan residues in protein molecule. The interaction of С5S and Au@C5S with BSA leads to the changes in the protein structure as well as in the physicochemical characteristics of gold nanoparticles, which is a promising aspect in the use of such systems for transport of protein molecules or their visualization-detection in biological media.

Tracking the effect of binder length on colloidal stability and bioconjugation of gold nanoparticles

Understanding the organization of self-assembled monolayers (SAMs) on gold nanoparticles (AuNPs) as protective coatings is a key role of biological applications of nanomaterials. Here, we report the influence on the stability of the surface coverage of three mercaptocarboxylic lingands onto AuNPs, mercaptopropanoic acid (MPA), mercaptoundecanoic acid (MUA) and mercaptopropionic acid (MHA) under different conditions. In addition, we optimized a bioconjugation route using bovine serum protein (BSA) as a protein model. AuNPs and successful binding of ligands and BSA on the AuNPs were analyzed by UV–Vis, TEM, FTIR, RAMAN, DLS and zeta potential. The size of as-synthesized AuNPs was 18 ± 1,2 nm with surface plasmon resonance (SPR) peak at 522 nm. The magnitude of the bathochromic shift of AuNPs with MPA, MUA and MHA was determined by UV–Vis and the SPR band position of AuNP shifts to 1.5, 3 and 5 nm longer. Moreover, the data show the influence of chain length on colloidal stability and covalent and non-covalent coupling steps with nanomaterials. We demonstrate a method for quantitative determination of the coatings on gold nanoparticles and open new perspectives in understanding the influence of monolayer thickness on the generation of nanobioconjugates for biological applications.

Characterization of the complex between native and reduced bovine serum albumin with aquacobalamin and evidence of dual tetrapyrrole binding.

Serum albumin binds to a variety of endogenous ligands and drugs. Human serum albumin (HSA) binds to heme via hydrophobic interactions and axial coordination of the iron center by protein residue Tyr161. Human serum albumin binds to another tetrapyrrole, cobalamin (Cbl), but the structural and functional properties of this complex are poorly understood. Herein, we investigate the reaction between aquacobalamin (H2OCbl) and bovine serum albumin (BSA, the bovine counterpart of HSA) using Ultraviolet-Visible and fluorescent spectroscopy, and electron paramagnetic resonance. The reaction between H2OCbl and BSA led to the formation of a BSA-Cbl(III) complex consistent with N-axial ligation (amino). Prior tothe formation of this complex, the reactants participate in an additional binding event that has been examined by fluorescence spectroscopy. Binding of BSA to Cbl(III) reduced complex formation between the bound cobalamin and free cyanide to form cyanocobalamin (CNCbl), suggesting that the β-axial position of the cobalamin may be occupied by an amino acid residue from the protein. Reaction of BSA containing reduced disulfide bonds with H2OCbl produces cob(II)alamin and disulfide with intermediate formation of thiolate Cbl(III)-BSA complex and its decomposition. Finally, in vitro studies showed that cobalamin binds to BSA only in the presence of an excess of protein, which is in contrast to heme binding to BSA that involves a 1:1 stoichiometry. In vitro formation of BSA-Cbl(III) complex does not preclude subsequent heme binding, which occurs without displacement of H2OCbl bound to BSA. These data suggest that the two tetrapyrroles interact with BSA in different binding pockets.

Pickering high internal phase emulsions stabilized by protein-covered cellulose nanocrystals

The development of high internal phase emulsions (HIPEs) stabilized by food-grade particles (e.g., cellulose nanocrystals, CNCs) has attracted increasing interests in the food colloid field. This paper reports a type of oil-in-water HIPEs using protein (bovine serum albumin, BSA)-covered CNCs as effective stabilizers. The BSA-covered CNCs were simply obtained by mixing aqueous solutions of anionic CNCs with the cationic BSA at pH 3.0. Turbidity and absorption measurements proved the binding of BSA to CNCs. The strong interaction and aggregation of BSA-covered CNCs were verified by atomic force microscopy measurements. Interestingly, the 0.5% (w/v) CNCs covered with a low amount of BSA (0.01%–0.1%, w/v) as the emulsifying agent yielded very stable, gel-like HIPEs (ϕ = 0.8). The stiffness and microstructure of the HIPEs can be modulated either by changing the surface protein coverage at a fixed CNC concentration or by changing the concentration of CNCs with a fixed surface protein coverage (e.g., 80%). The improvement of the stiffness was largely attributed to the enhanced droplet packing effect and interfacial bridging effect. These findings offer a promising opportunity to extend the applications of protein-modified CNCs in the food and pharmaceutical industries.

Investigating the structural integrity of Bovine serum albumin in presence of newly synthesized metallosurfactants

Bovine Serum Albumin is major transport protein and is often used as a drug carrier in body organs. Knowledge of its binding with metallosurfactant can significantly influence the biodistribution of metallodrugs. Current work demonstrated a facile method to prepare four different double chained metallosurfactants containing Fe, Co, Ni and Cu as part of their counter ion. The as-synthesized metallosurfactants were characterized using FTIR, AAS, TGA and XRD in solid form. The aggregation of these metallosurfactants in aqueous medium was investigated through conductivity, surface tension and SAXS. Further, we have investigated their binding with BSA through different analytical methods The effect of concentration of metallosurfactants on the primary and secondary structure of BSA was further examined by SDS-PAGE and Circular dichroism, respetively. It is found that at premicellar concentration, the primary structure of BSA was not affected but the secondary structure i.e. α-helical structure of BSA was altered as shown by circular dichroism. Interestingly, post micellar concentration of metallosurfactants shows the pronounced effect on the primary and secondary structure of BSA. SAXS study also supports the fact of unfolding of protein and its wrapping around the micelles. Zeta potential describes the electrical charge and stability of the protein in the presence of different concentration of metallosurfactant. Along with, it was found that presence of protein delays the aggregation behavior of metallosurfactant, as a sign of binding of BSA with metallosurfactant.

Synergistic effect of chitosan-zinc-tin oxide colloidal nanoparticle and their binding performance on bovine albumin serum

We report the hydrophobic interaction of chitosan-zinc-tin oxide nanoparticle with bovine albumin serum (BSA). A new renewable material of chitosan (N-deacetylated derivative of chitin) was subjected to action with zinc oxide and tin oxide modifiers. The nanoparticles of chitosan-zinc-tin oxide have been successfully synthesized by simple chemical precipitation method. The synthesized nanoparticles were characterized and confirmed by FTIR, XRD, HR-SEM with EDAX, TEM and BET analysis. This result clearly indicates that the colloidal nanoparticles are quenches the fluorescence of BSA through the ground state complex formation via static quenching mechanism. The quenching rate constant, binding sites, binding force and binding constant (K) = 0.80–1.41 × 104 M−1 were calculated from fluorescence spectral data and shows that the hydrophobic interaction is observed between colloidal nanoparticle and BSA. Thus the efficient binding of BSA with colloidal chitosan-zinc-tin oxide nanoparticle leads to play an important role in biomedical, biosensors and pharmaceutical applications.

Probing the binding behavior and kinetics of silver nanoparticles with bovine serum albumin

The binding of BSA to AgNPs will open a door to fully understand the essence of the interactions between nanoparticles and proteins.

Effects of serum albumin on the degradation and cytotoxicity of single-walled carbon nanotubes

Neutrophil myeloperoxidase (MPO) and peroxynitrite (ONOO−) can oxidatively biodegrade carboxylated single-walled carbon nanotubes (SWCNTs). The protein–SWCNTs interactions will play an important role in the degradation and cytotoxicity of nanotubes. Here, we investigated the binding of bovine serum albumin (BSA, a common and well-characterized model blood serum protein) to SWCNTs, and found that the hydrophobic and electrostatic interactions might be crucial factors in stabilizing the binding of SWCNTs with BSA. The binding of BSA could impair SWCNTs biodegradation in vitro through the competitive adsorption to nanotube. Both SWCNTs and BSA-SWCNTs were significantly degraded in zymosan-stimulated macrophages, and the degradation degree was more for BSA-SWCNTs. The mechanism for SWCNTs degradation in activated macrophages was further investigated to demonstrate the dominant participation of MPO and ONOO−-driven pathways. Moreover, binding of BSA to SWCNTs reduced cytotoxicity and degraded nanotubes induced less cytotoxicity than non-degraded nanotubes. The binding of BSA may be an important determinant for the biodegradation and cytotoxicity of SWCNTs in inflammatory cells, and therefore, provide a new route to mitigate the potential toxicity of nanotubes in future biomedical applications.

Recognition of Malondialdehyde-modified Proteins by the C Terminus of Complement Factor H Is Mediated via the Polyanion Binding Site and Impaired by Mutations Found in Atypical Hemolytic Uremic Syndrome

Atypical hemolytic uremic syndrome (aHUS) is a severe thrombotic microangiopathy characterized by uncontrolled complement activation against endothelial and blood cells. Mutations in the C-terminal target recognition domains 19-20 of complement regulator factor H (FH) are strongly associated with aHUS, but the mechanisms triggering disease onset have remained unresolved. Here we report that several aHUS-related mutations alter the binding of FH19-20 to proteins where lysines have reacted with malondialdehyde (MDA). Although FH19-20 did not interact with MDA-modified hexylamine, lysine-containing peptides, or a proteolytically degraded protein, it bound to MDA-modified polylysine. This suggests that FH19-20 recognizes only clustered MDA adducts. Binding of MDA-modified BSA to FH19-20 was ionic by nature, depended on positive residues of FH19-20, and competed with the polyanions heparin and DNA. This could not be explained with the mainly neutral adducts known to form in MDA modification. When positive charges of lysines were eliminated by acetic anhydride instead of MDA, the acetylated BSA started to bind FH19-20. Together, these results indicate that negative charges on the modified proteins dominate the interaction with FH19-20. This is beneficial for the physiological function of FH because by binding to the negative charges of the modified target, FH could prevent excess complement activation initiated by naturally occurring antibodies recognizing MDA epitopes with multiple different structures. We propose that oxidative stress leading to formation of MDA adducts is a common feature for triggers of aHUS and that failure of FH in protecting MDA-modified surfaces from complement activation is involved in the pathogenesis of the disease.

Tuning the Activities and Structures of Enzymes Bound to Graphene Oxide with a Protein Glue

Graphene oxide (GO) is being investigated extensively for enzyme and protein binding, but many enzymes bound to GO denature considerably and lose most of their activities. A simple, novel, and efficient approach is described here for improving the structures and activities of enzymes bound to GO such that bound enzymes are nearly as active as those of the corresponding unbound enzymes. Our strategy is to preadsorb highly cationized bovine serum albumin (cBSA) to passivate GO, and cBSA/GO (bGO) served as an excellent platform for enzyme binding. The binding of met-hemoglobin, glucose oxidase, horseradish peroxidase, BSA, catalase, lysozyme, and cytochrome c indicated improved binding, structure retention, and activities. Nearly 100% of native-like structures of all the seven proteins/enzymes were noted at near monolayer formation of cBSA on GO (400% w/w), and all bound enzymes indicated 100% retention of their activities. A facile, benign, simple, and general method has been developed for the biofunctionalization of GO, and this approach of coating with suitable protein glues expands the utility of GO as an advanced biophilic nanomaterial for applications in catalysis, sensing, and biomedicine.

Most Methylation-Susceptible DNA Sequences in Human Embryonic Stem Cells Undergo a Change in Conformation or Flexibility upon Methylation

DNA methylation in eukaryotes occurs on the cytosine bases in CG, CHG, and CHH (where H indicates non-G nucleotides) contexts and provides an important epigenetic mark in various biological processes. However, the structural and physical properties of methylated DNA are poorly understood. Using nondenaturing polyacrylamide gel electrophoresis, we performed a systematic study of the influence of DNA methylation on the conformation and physical properties of DNA for all CG, CHG, and CHH contexts. In the CG context, methylated multimers of the CG/CG-containing unit fragment migrated in gels slightly faster than their unmethylated counterparts. In the CHG context, both homo- and hemimethylation caused retarded migration of multimers of the CAG/CTG-containing fragment. In the CHH context, methylation caused or enhanced retarded migration of the multimers of CAA/TTG-, CAT/ATG-, CAC/GTG-, CTA/TAG-, or CTT/AAG-containing fragments. These results suggest that methylation increases DNA rigidity in the CG context and introduces distortions into several CHG and CHH sequences. More interestingly, we found that nearly all of the methylation repertoires in the CHG context and 98% of those in the CHH context in human embryonic stem cells were species that undergo conformational changes upon methylation. Similarly, most of the methylation repertoires in the Arabidopsis CHG and CHH contexts were sequences with methylation-induced distortion. We hypothesize that the methylation-induced properties or conformational changes in DNA may facilitate nucleosome formation, which provides the essential mechanism for alterations of chromatin density.

Effect of protein thermo aggregation on the binding of BSA to gelatin type A

We study the effect of limited heat-induced aggregation of BSA on structure development in the water–gelatin-thermally aggregated BSA (BSATA) system. The pH is set at 5.4 and the temperature is higher than the conformation transition temperature of gelatin, but lower than the denaturation temperature of BSA. Dynamic light scattering, circular dichroism, and fluorescence measurements are used to monitor structure changes. Interaction of gelatin with BSATA leads to formation of large complex particles with an average radius ∼1500nm. BSA–gelatin complex formation accompanies partial destabilization of the secondary and tertiary structures of BSA and an additional exposure of hydrophobic tryptophan residues on the surface of the globule. It is shown that electrostatic interaction of the oppositely charged groups of BSATA and gelatin is responsible for formation of such complex particles, whereas the secondary forces (hydrophobic interaction and hydrogen bonds) play an important role in stabilization of the complex particles. The zeta potentials of the native and the thermally aggregated BSA samples were determined, and the solvent quality has been quantified by determining the activity of the protein samples in their saturated solutions. It was shown that steric reasons (large size of the thermally aggregated BSATA particles), and uncomplete charge compensation of the positively charged gelatin molecules by the negatively charged BSATA particles are the main factors in determining structure formation, while the levels of the activity of the native BSA and BSATA have a smaller effect on the structure of complex.

Tris(hydroxymethyl)aminomethane‐functionalized agarose particles: Parameters affecting the binding of bovine serum albumin

A new protein adsorbent is introduced based on the coupling of the common buffer ion, tris(hydroxymethyl)aminomethane, to the agarose gel Sepharose HP from GE Healthcare Bio-Sciences AB, Uppsala, Sweden. The article describes the synthesis of the new adsorbent and the use of BSA as a model in a binding study. By optimization of the coupling procedure, a maximum ligand density of 63.5 μmol/mL gel could be obtained. Adsorption equilibria were investigated in the pH range 5.0-8.0 and at salt concentrations of 0-0.4 mol/L. Binding of BSA under different conditions indicated that both electrostatic interaction and hydrogen bonding were involved in the adsorption process where the former played a major role.

Albumin-Binding and Angiogenic Domains of SPARC Located at Its C-Terminus.

Abstract Background: SPARC (Secreted Protein Acidic and Rich in Cysteine) is an albumin-binding glycoprotein overexpressed in breast cancer and is associated with poor prognosis and shorter overall survival. We have previously shown that SPARC expression appeared to correlate with response to nanoparticle albumin-bound (nab) paclitaxel (Abraxane) in head-and-neck and pancreatic-cancer patients. To further define the role of SPARC, recombinant-human SPARC (rhSPARC) was characterized to identify its albumin-binding and angiogenic domain.Materials and methods: The albumin-binding site on SPARC was defined by testing the binding of Alexa 488-labeled BSA to immobilized rhSPARC in a solid-phase-binding assay and in a competitive-binding assay in presence of increasing concentration of various SPARC-derived peptides. The angiogenic activity of rhSPARC was evaluated using a HUVEC tube-formation assay.Results: The SPARC albumin-binding assay revealed a pattern of saturable and specific binding with an estimated Kd of 700 µM very near the known plasma concentration of albumin (600 µM). Competitive binding with Cathepsin K-digested SPARC and SPARC-derived peptides defined the albumin-binding domain to be amino acids 209-223 in the C-terminal region. In the HUVEC tube-formation assay, rhSPARC was pro-angiogenic at 10 µg/ml and anti-angiogenic at 100 µg/ml. The angiogenic domain of SPARC was located to the C-terminal of SPARC.Discussion: The albumin-binding domain of SPARC was located to residues aa209-223, supporting its role as a target for nab-technology-based drug delivery. We confirmed that SPARC can promote angiogenesis at physiological concentrations, contributing to the role of SPARC in a more aggressive tumor phenotype. The angiogenic domain of SPARC is located within a 54-aa sequence of its C-terminus. SPARC could be a potential therapeutic target and a biomarker for predicting response to nab-paclitaxel. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 2144.