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At least 451 records · Page 25
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Causal relationship between mitochondrial-associated proteins and cerebral aneurysms: a Mendelian randomization study

Background Cerebral aneurysm is a high-risk cerebrovascular disease with a poor prognosis, potentially linked to multiple factors. This study aims to explore the association between mitochondrial-associated proteins and the risk of cerebral aneurysms using Mendelian randomization (MR) methods. Methods We used GWAS summary statistics from the IEU Open GWAS project for mitochondrial-associated proteins and from the Finnish database for cerebral aneurysms (uIA, aSAH). The association between mitochondrial-associated exposures and cerebral aneurysms was evaluated using MR-Egger, weighted mode, IVW, simple mode and weighted median methods. Reverse MR assessed reverse causal relationship, while sensitivity analyses examined heterogeneity and pleiotropy in the instrumental variables. Significant causal relationship with cerebral aneurysms were confirmed using FDR correction. Results Through MR analysis, we identified six mitochondrial proteins associated with an increased risk of aSAH: AIF1 (OR: 1.394, 95% CI: 1.109–1.752, p = 0.0044), CCDC90B (OR: 1.318, 95% CI: 1.132–1.535, p = 0.0004), TIM14 (OR: 1.272, 95% CI: 1.041–1.553, p = 0.0186), NAGS (OR: 1.219, 95% CI: 1.008–1.475, p = 0.041), tRNA PusA (OR: 1.311, 95% CI: 1.096–1.569, p = 0.003), and MRM3 (OR: 1.097, 95% CI: 1.016–1.185, p = 0.0175). Among these, CCDC90B, tRNA PusA, and AIF1 demonstrated a significant causal relationship with an increased risk of aSAH (FDR q < 0.1). Three mitochondrial proteins were associated with an increased risk of uIA: CCDC90B (OR: 1.309, 95% CI: 1.05–1.632, p = 0.0165), tRNA PusA (OR: 1.306, 95% CI: 1.007–1.694, p = 0.0438), and MRM3 (OR: 1.13, 95% CI: 1.012–1.263, p = 0.0303). In the reverse MR study, only one mitochondrial protein, TIM14 (OR: 1.087, 95% CI: 1.004–1.177, p = 0.04), showed a causal relationship with aSAH. Sensitivity analysis did not reveal heterogeneity or pleiotropy. The results suggest that CCDC90B, tRNA PusA, and MRM3 may be common risk factors for cerebral aneurysms (ruptured and unruptured), while AIF1 and NAGS are specifically associated with an increased risk of aSAH, unrelated to uIA. TIM14 may interact with aSAH. Conclusion Our findings confirm a causal relationship between mitochondrial-associated proteins and cerebral aneurysms, offering new insights for future research into the pathogenesis and treatment of this condition.

Wang, Shuai↗
DOE OSTI2024

Hepatic transcriptome analyses of juvenile white bass ( Morone chrysops ) when fed diets where fish meal is partially or totally replaced by alternative protein sources

White bass (Morone chrysops) are a popular sportfish throughout the southern United States, and one parent of the commercially-successful hybrid striped bass (M. chrysops male x M. saxatilis female). Currently, white bass are cultured using diets formulated for other carnivorous fish, such as largemouth bass (Micropterus salmoides) or hybrid striped bass and contain a significant percentage of marine fish meal. Since there are no studies regarding the utilization of alternative proteins in this species, we evaluated the global gene expression of white bass fed diets in which fish meal was partially or totally replaced by various combinations of soybean meal, poultry by-product meal, canola meal, soy protein concentrate, wheat gluten, or a commercial protein blend (Pro Cision™). Six isonitrogenous (40% protein), isolipidic (11%), and isocaloric (17.1 kJ/g) diets were formulated to meet the known nutrient and energy requirements of largemouth bass and hybrid striped bass using nutrient availability data for most of the dietary ingredients. One of the test diets consisted exclusively of plant protein sources. Juvenile white bass (40.2 g initial weight) were stocked into a flow-through aquaculture system (three tanks/diet; 10 fish/tank) and fed the test diets twice daily to satiation for 60 days. RNA sequencing and bioinformatic analyses revealed significant differentially expressed genes between all test diets when compared to fish meal control. A total of 1,260 differentially expressed genes were identified, with major ontology relating to cell cycle and metabolic processes as well as immune gene functions. This data will be useful as a resource for future refinements to moronid diet formulation, as marine fish meal becomes limiting and plant ingredients are increasingly added as a reliable protein source

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

The Promise of Mutation Resistant Drugs for SARS-CoV-2 That Interdict in the Folding of the Spike Protein Receptor Binding Domain

In recent work, we proposed that effective therapeutic drugs aimed at treating the SARS-CoV-2 infection could be developed based on interdicting in the early steps of the folding pathway of key viral proteins, including the receptor binding domain (RBD) of the spike protein. In order to provide for a drug target on the protein, the earliest contact-formation event along the dominant folding pathway of the RBD spike protein was predicted employing the Sequential Collapse Model (SCM). The segments involved in the predicted earliest contact were suggested to provide optimal folding interdiction target regions (FITRs) for potential therapeutic drugs, with a focus on folding interdicting peptides (FIPs). In this paper, we extend our analysis to include 13 known single mutations of the RBD spike protein as well as the triple mutation B1.351 and the recent double mutation B1.617.2. The results show that the location of the FITR does not change in any of the 15 studied mutations, providing for a mutation-resistant drug design strategy for the RBD-spike protein.

Bergasa-Caceres, Fernando↗
DOE OSTI2021

Bioinformatics Investigations of Universal Stress Proteins from Mercury-Methylating Desulfovibrionaceae

The presence of methylmercury in aquatic environments and marine food sources is of global concern. The chemical reaction for the addition of a methyl group to inorganic mercury occurs in diverse bacterial taxonomic groups including the Gram-negative, sulfate-reducing Desulfovibrionaceae family that inhabit extreme aquatic environments. The availability of whole-genome sequence datasets for members of the Desulfovibrionaceae presents opportunities to understand the microbial mechanisms that contribute to methylmercury production in extreme aquatic environments. We have applied bioinformatics resources and developed visual analytics resources to categorize a collection of 719 putative universal stress protein (USP) sequences predicted from 93 genomes of Desulfovibrionaceae. We have focused our bioinformatics investigations on protein sequence analytics by developing interactive visualizations to categorize Desulfovibrionaceae universal stress proteins by protein domain composition and functionally important amino acids. We identified 651 Desulfovibrionaceae universal stress protein sequences, of which 488 sequences had only one USP domain and 163 had two USP domains. The 488 single USP domain sequences were further categorized into 340 sequences with ATP-binding motif and 148 sequences without ATP-binding motif. The 163 double USP domain sequences were categorized into (1) both USP domains with ATP-binding motif (3 sequences); (2) both USP domains without ATP-binding motif (138 sequences); and (3) one USP domain with ATP-binding motif (21 sequences). We developed visual analytics resources to facilitate the investigation of these categories of datasets in the presence or absence of the mercury-methylating gene pair (hgcAB). Future research could utilize these functional categories to investigate the participation of universal stress proteins in the bacterial cellular uptake of inorganic mercury and methylmercury production, especially in anaerobic aquatic environments.

Isokpehi, Raphael D. (ORCID:0000000268770840)↗
DOE OSTI2021

NMR Spectroscopy for Protein Higher Order Structure Similarity Assessment in Formulated Drug Products

Peptide and protein drug molecules fold into higher order structures (HOS) in formulation and these folded structures are often critical for drug efficacy and safety. Generic or biosimilar drug products (DPs) need to show similar HOS to the reference product. The solution NMR spectroscopy is a non-invasive, chemically and structurally specific analytical method that is ideal for characterizing protein therapeutics in formulation. However, only limited NMR studies have been performed directly on marketed DPs and questions remain on how to quantitively define similarity. Here, NMR spectra were collected on marketed peptide and protein DPs, including calcitonin-salmon, liraglutide, teriparatide, exenatide, insulin glargine and rituximab. The 1D 1 H spectral pattern readily revealed protein HOS heterogeneity, exchange and oligomerization in the different formulations. Principal component analysis (PCA) applied to two rituximab DPs showed consistent results with the previously demonstrated similarity metrics of Mahalanobis distance (D M ) of 3.3. The 2D 1 H- 13 C HSQC spectral comparison of insulin glargine DPs provided similarity metrics for chemical shift difference (Δδ) and methyl peak profile, i.e., 4 ppb for 1 H, 15 ppb for 13 C and 98% peaks with equivalent peak height. Finally, 2D 1 H- 15 N sofast HMQC was demonstrated as a sensitive method for comparison of small protein HOS. The application of NMR procedures and chemometric analysis on therapeutic proteins offer quantitative similarity assessments of DPs with practically achievable similarity metrics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Swine Host Protein Coiled-Coil Domain-Containing 115 (CCDC115) Interacts with Classical Swine Fever Virus Structural Glycoprotein E2 during Virus Replication

Interactions between the major structural glycoprotein E2 of classical swine fever virus (CSFV) with host proteins have been identified as important factors affecting virus replication and virulence. Previously, using the yeast two-hybrid system, we identified swine host proteins specifically interacting with CSFV E2. In this report, we use a proximity ligation assay to demonstrate that swine host protein CCDC115 interacts with E2 in CSFV-infected swine cells. Using a randomly mutated E2 library in the context of a yeast two-hybrid methodology, specific amino acid mutations in the CSFV E2 protein responsible for disrupting the interaction with CCDC115 were identified. A recombinant CSFV mutant (E2ΔCCDC115v) harboring amino acid changes disrupting the E2 protein interaction with CCDC115 was produced and used as a tool to assess the role of the E2–CCDC115 interaction in viral replication and virulence in swine. CSFV E2ΔCCDC115v showed a slightly decreased ability to replicate in the SK6 swine cell line and a greater replication defect in primary swine macrophage cultures. A decreased E2–CCDC115 interaction detected by PLA is observed in cells infected with E2ΔCCDC115v. Importantly, animals intranasally infected with 105 TCID50 of E2ΔCCDC115v experienced a significantly longer survival period when compared with those infected with the parental Brescia strain. This result would indicate that the ability of CSFV E2 to bind host CCDC115 protein during infection plays an important role in virus replication in swine macrophages and in virus virulence during the infection in domestic swine.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Protein Molecular Dynamics Simulations with Approximate QM: What Can We Learn?

Classical force fields are essential for computer simulations of proteins and are typically parameterized to reproduce secondary and tertiary structure of isolated proteins. However, while protein-protein interactions are ubiquitous in nature, they are not considered in parameterization efforts and are far less understood than isolated proteins. A better characterization of intermolecular interactions is widely recognized as a key to revolutionizing drug and therapeutic developments with high-throughput computational screening. Urgently needed is a critical assessment of the performance of modern protein force fields against first-principles electronic structure methods and experiments. In a daring step toward this goal, we here describe a comparison of peptide folding dynamics as predicted by a molecular mechanics force field on the one hand and by an approximate electronic structure quantum mechanical (QM) method based on density-functional tight-binding (DFTB) on the other. We further compare the dynamics from straightforward DFTB simulations with a near-linear scaling version of DFTB for massively parallel computation based on the fragment molecular orbital (FMO-DFTB) method. We illustrate differences between the phenomenology of the folding dynamics from these three methods for a small model peptide, as well as charge polarization and dynamic fluctuations, point out possible correlations and implications for force field developers, and discuss the lessons learned that might become applicable to future predictive high-throughput computer screening for personalized neoantigen cancer therapy.

Irle, Stephan↗
DOE OSTI2025

Smart culture medium optimization for recombinant protein production: Experimental, modeling, and AI/ML-driven strategies

Recombinant protein production (RPP) is central to biotechnology, where recombinant proteins are used as either end products or catalysts in the synthesis of chemicals, fuels, and materials. Among the major cost drivers, culture medium plays a pivotal role in determining protein yield and quality. This review presents a comprehensive perspective on the critical stages of “smart” culture medium optimization: planning, screening, modeling, optimization, and validation. In the planning stage, we examine the nutritional and energetic roles of medium components, including carbon, nitrogen, amino acids, salts, and trace metals, and their impacts on culture parameters such as pH, oxidative state, and osmolality. We highlight the variability in trace metal content due to water sources, culture vessels, and raw materials, which can substantially influence RPP. The screening stage covers Design of Experiments (DoE) approaches, assessing their theoretical basis, implementation, and limitations. For modeling, we describe methods that integrate experimental data to develop predictive models for smart medium formulation. Model-based optimization strategies can then be employed to select optimal media compositions for a given application. The validation stage aims to evaluate model predictions and provide feedback for model training and refinement. Finally, we survey mechanistic and artificial intelligence/machine learning (AI/ML)-driven models as integrated, transformational tools for predictive modeling of bioprocess conditions, nutrient availability, cellular metabolism, and protein quality, with the goal of optimizing culture media to enhance protein yields while reducing costs and environmental impact. We conclude by addressing the challenges of translating laboratory-scale medium optimization to industrial-scale settings and exploring future AI/ML-driven approaches that may overcome current bottlenecks and accelerate medium design for RPP. Overall, this review provides a unified framework for advancing smart medium design in RPP.

Artificial Intelligence/Machine Learning (AI/ML)↗
DOE OSTI2024

Orange carotenoid proteins: structural understanding of evolution and function

Cyanobacteria uniquely contain a primitive water-soluble carotenoprotein, the orange carotenoid protein (OCP). Nearly all extant cyanobacterial genomes contain genes for the OCP or its homologs, implying an evolutionary constraint for cyanobacteria to conserve its function. Genes encoding the OCP and its two constituent structural domains, the N-terminal domain, helical carotenoid proteins (HCPs), and its C-terminal domain, are found in the most basal lineages of extant cyanobacteria. These three carotenoproteins exemplify the importance of the protein for carotenoid properties, including protein dynamics, in response to environmental changes in facilitating a photoresponse and energy quenching. Furthermore, we review new structural insights for these carotenoproteins and situate the role of the protein in what is currently understood about their functions.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Comparative Pore Structure and Dynamics for Bacterial Microcompartment Shell Protein Assemblies in Sheets or Shells

Bacterial microcompartments (BMCs) are protein-bound organelles found in some bacteria that encapsulate enzymes for enhanced catalytic activity. These compartments spatially sequester enzymes within semipermeable shell proteins, analogous to many membrane-bound organelles. The shell proteins assemble into multimeric tiles; hexamers, trimers, and pentamers, and these tiles self-assemble into larger assemblies with icosahedral symmetry. While icosahedral shells are the predominant form in vivo , the tiles can also form nanoscale cylinders or sheets. The individual multimeric tiles feature central pores that are key to regulating transport across the protein shell. Our primary interest is to quantify pore shape changes in response to alternative component morphologies at the nanoscale. We used molecular modeling tools to develop atomically detailed models for both planar sheets of tiles and curved structures representative of the complete shells found in vivo . Subsequently, these models were animated using classical molecular dynamics simulations. From the resulting trajectories, we analyzed the overall structural stability, water accessibility to individual residues, water residence time, and pore geometry for the hexameric and trimeric protein tiles from the Haliangium ochraceu m model BMC shell. These exhaustive analyses suggest no substantial variation in pore structure or solvent accessibility between the flat and curved shell geometries. We additionally compare our analysis to hydroxyl radical footprinting data to serve as a check against our simulation results, highlighting specific residues where water molecules are bound for a long time. Although with little variation in morphology or water interaction, we propose that the planar and capsular morphology can be used interchangeably when studying permeability through BMC pores.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2025

Bond-centric modular design of protein assemblies

Directional interactions that generate regular coordination geometries are a powerful means of guiding molecular and colloidal self-assembly, but implementing such high-level interactions with proteins remains challenging due to their complex shapes and intricate interface properties. Here we describe a modular approach to protein nanomaterial design inspired by the rich chemical diversity that can be generated from the small number of atomic valencies. We design protein building blocks using deep learning-based generative tools, incorporating regular coordination geometries and tailorable bonding interactions that enable the assembly of diverse closed and open architectures guided by simple geometric principles. Experimental characterization confirms the successful formation of more than 20 multicomponent polyhedral protein cages, two-dimensional arrays and three-dimensional protein lattices, with a high (10%–50%) success rate and electron microscopy data closely matching the corresponding design models. Due to modularity, individual building blocks can assemble with different partners to generate distinct regular assemblies, resulting in an economy of parts and enabling the construction of reconfigurable networks for designer nanomaterials.

Biomaterials – proteins↗
DOE OSTI2024

De novo design of buttressed loops for sculpting protein functions

In natural proteins, structured loops have central roles in molecular recognition, signal transduction and enzyme catalysis. However, because of the intrinsic flexibility and irregularity of loop regions, organizing multiple structured loops at protein functional sites has been very difficult to achieve by de novo protein design. Here we describe a solution to this problem that designs tandem repeat proteins with structured loops (9–14 residues) buttressed by extensive hydrogen bonding interactions. Experimental characterization shows that the designs are monodisperse, highly soluble, folded and thermally stable. Crystal structures are in close agreement with the design models, with the loops structured and buttressed as designed. We demonstrate the functionality afforded by loop buttressing by designing and characterizing binders for extended peptides in which the loops form one side of an extended binding pocket. The ability to design multiple structured loops should contribute generally to efforts to design new protein functions.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2021

Purification of native histidine-rich protein 2 (nHRP2) from Plasmodium falciparum culture supernatant, infected RBCs, and parasite lysate

Background: Despite the widespread use of histidine-rich protein 2 (HRP2)-based rapid diagnostic tests (RDTs), purified native HRP2 antigen is not standardly used in research applications or assessment of RDTs used in the field. Methods: This report describes the purification of native HRP2 (nHRP2) from the HB3 Plasmodium falciparum culture strain. As this culture strain lacks pfhrp3 from its genome, it is an excellent source of HRP2 protein only and does not produce the closely-related HRP3. The nHRP2 protein was isolated from culture supernatant, infected red blood cells (iRBCs), and whole parasite lysate using nickel-metal chelate chromatography. Biochemical characterization of nHRP2 from HB3 culture was conducted by SDS-PAGE and western blotting, and nHRP2 was assayed by RDT, ELISA, and bead-based immunoassay. Results: Purified nHRP2 was identified by SDS-PAGE and western blot as a –60 kDa protein that bound anti-HRP-2 monoclonal antibodies. Mouse anti-HRP2 monoclonal antibody was found to produce high optical density readings between dilutions of 1:100 and 1:3,200 by ELISA with assay signal observed up to a 1:200,000 dilution. nHRP2 yield from HB3 culture by bead-based immunoassay revealed that both culture supernatant and iRBC lysate were practical sources of large quantities of this antigen, producing a total yield of 292.4 µg of nHRP2 from two pooled culture preparations. Assessment of nHRP2 recognition by RDTs revealed that Carestart Pf HRP2 and HRP2/pLDH RDTs detected purified nHRP2 when applied at concentrations between 20.6 and 2060 ng/mL, performing within a log-fold dilution of commercially-available recombinant HRP2. The band intensity observed for the nHRP2 dilutions was equivalent to that observed for P. falciparum culture strain dilutions of 3D7 and US06 F Nigeria XII between 12.5 and 1000 parasites/µL. Conclusions: Purified nHRP2 could be a valuable reagent for laboratory applications as well as assessment of new and existing RDTs prior to their use in clinical settings. These results establish that it is possible to extract microgram quantities of the native HRP2 antigen from HB3 culture and that this purified protein is well recognized by existing monoclonal antibody lines and RDTs.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Acute total body ionizing gamma radiation induces long-term adverse effects and immediate changes in cardiac protein oxidative carbonylation in the rat

Radiation-induced heart disease presents a significant challenge in the event of an accidental radiation exposure as well as to cancer patients who receive acute doses of irradiation as part of radiation therapy. We utilized the spontaneously hypertensive Wistar-Kyoto rat model, previously shown to demonstrate drug-induced cardiomyopathy, to evaluate the acute and long-term effects of sub-lethal total body gamma irradiation at two, four, and fiftytwo weeks. We further examined irreversible oxidative protein carbonylation in the heart immediately following irradiation in the normotensive Wistar-Kyoto rat. Both males and females sustained weight loss and anemic conditions compared to untreated controls over a one-year period as reflected by reduced body weight and low red blood cell count. Increased inflammation was detected by elevated IL-6 serum levels selectively in males at four weeks. Serum cardiac troponin T and I analyses revealed signs of cardiomyopathy at earlier timepoints, but high variability was observed, especially at one year. Echocardiography at two weeks following 5.0Gy treatment revealed a significant decrease in cardiac output in females and a significant decrease in both diastolic and systolic volumes in males. Following 10.0Gy irradiation in the normotensive Wistar-Kyoto rat, the heart tissue showed an increase in total protein oxidative carbonylation accompanied by DNA damage indicated by an increase in γ-H2AX. Using proteomic analyses, we identified several novel proteins which showed a marked difference in carbonylation including those of mitochondrial origin and most notably, cardiac troponin T, one of the key proteins involved in cardiomyocyte contractility. Overall, we present findings of acute oxidative protein damage, DNA damage, cardiac troponin T carbonylation, and long-term cardiomyopathy in the irradiated animals.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Probing lanmodulin's mechanisms of rare-earth selectivity for protein-based bioseparations

Our BES Separation Science program project, DE-SC0021007, supported our efforts to begin to understand the mechanisms underlying selectivity of a novel class of lanthanide-binding proteins discovered by our laboratory, called lanmodulin (LanM), and to leverage these proteins for recovery and separations of trivalent rare earth elements (REEs) as well as of trivalent actinides. Overall, our work provides important insights into how higher-order (e.g., secondary, tertiary, and quaternary) protein structure modulates selectivity profiles of proteins that bind f-elements highly selectively. These results are important for advancing the concept of protein-based separations of REEs and, perhaps, of other critical minerals.

Lanmodulin, rare earth elements, protein-based met↗
DOE OSTI2023

Deep Green Unannotated Protein Structures

The Deep Green list is based on the identification and curation of conserved unannotated proteins in three green lineage (Viridiplantae) model organisms; Arabidopsis thaliana, Chlamydomonas reinhardtii, and Setaria viridis. Preliminary characterization of Deep Green proteins and genes was done using various informatics tools and published data sets and is presented in Knoshaug, Sun, et al., 2023, submitted. The structures of these unannotated proteins were also predicted using AlphaFold (Jumper et al., 2021). The data deposited here are the AlphaFold structural predictions having the highest pLDDT score and thus identified as the best folded structure (ranked_0). These data enable others to do in-depth structural characterizations to aid in functional characterization leading to deeper understanding of plant biology. References: Jumper, J., Evans, R., Pritzel, A., Green, T., Figurnov, M., Ronneberger, O., Tunyasuvunakool, K., Bates, R., Žídek, A., Potapenko, A., Bridgland, A., Meyer, C., Kohl, S. A. A., Ballard, A. J., Cowie, A., Romera-Paredes, B., Nikolov, S., Jain, R., Adler, J., Back, T., Petersen, S., Reiman, D., Clancy, E., Zielinski, M., Steinegger, M., Pacholska, M., Berghammer, T., Bodenstein, S., Silver, D., Vinyals, O., Senior, A. W., Kavukcuoglu, K., Kohli, P. and Hassabis, D. (2021) Highly accurate protein structure prediction with AlphaFold. Nature, 596:583-589. Knoshaug, E. P., Sun, P., Nag, A., Nguyen, H., Mattoon, E. M., Zhang, N., Liu, J., Chen, C., Cheng, J., Zhang, R., St. John, P., and Umen, J. (submitted) Identification and preliminary characterization of conserved uncharacterized proteins from Chlamydomonas reinhardtii, Arabidopsis thaliana, and Setaria viridis.

09 BIOMASS FUELS↗
DOE OSTI2025

The Anisotropic Adsorption of De Novo Allosteric Two‐Component Protein Fibers on Mica Surfaces

Protein adsorption at solid–liquid interfaces underlies many biomedical and materials applications, yet the mechanisms governing adsorption of proteins and their assemblies remain poorly understood. Here we investigate de novo–designed proteins that self-assemble into three fibrillar morphologies — small (S), large (L), and helical (H)—on muscovite mica. S-fibers are metastable, forming first but diminishing as L- and H-fibers develop and deposit. Adsorption of both fibers and monomers depends on fiber morphology and solvent environment, which is modulated by the substrate. The anisotropic surface features of the fiber types—long axis of S- and L-fibers and helical grooves of H-fibers—correlate with specific crystallographic directions on mica (001). S- and L-fibers align along the unique lattice axis, while the substrate-facing groove of H-fibers preferentially aligns along the remaining symmetry-related directions. Increasing potassium chloride (KCl) concentrations to molar levels alters adsorption, decreasing monomer coverage relative to fibers. These observations, interpreted through the effect of interfacial interface, indicate that ion-mediated modulation of hydration layers governs electrostatic interactions and alignment. This study reveals how coupling between protein topography, substrate crystallography, and hydration structure dictates selective adsorption and orientation of protein assemblies, offering insights for rational biomolecular material design.

AFM↗
DOE OSTI2022

Predicting transport of intra-articularly injected growth factor fusion proteins into human knee joint cartilage

There are no drugs or treatment methods known to prevent the development of post-traumatic osteoarthritis (PTOA), a type of osteoarthritis (OA) that is triggered by traumatic joint injuries and accounts for ~12% of the nearly 600 million OA cases worldwide. Lack of effective drug delivery techniques remains a major challenge in developing clinically effective treatments, but cationic delivery carriers can help overcome this challenge. Scaling up treatments that are effective in in vitro models to achieve success in preclinical in vivo models and clinical trials is also a challenging problem in the field. Here we use a cationic green fluorescent protein (GFP) as a carrier to deliver Insulin-Like Growth Factor 1 (IGF-1), a drug considered as a potential therapeutic for PTOA. GFP-IGF-1 conjugates were first synthesized as fusion proteins with different polypeptide linkers, and their transport properties were characterized in human cartilage explants. In vitro experimental data were used to develop a predictive mathematical transport model that was validated using an independent in vitro experimental data set. In this study, the model was used to predict the transport of these fusion proteins upon intra-articular injection into human knee joints. The predictions included results for the rate and extent of fusion protein penetration into cartilage, and the maximum levels of fusion proteins that would escape into systemic circulation through the joint capsule. Together, our transport measurements and model set the stage for translation of such explant culture studies to in vivo preclinical studies and potentially clinical application.

36 MATERIALS SCIENCE↗