Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P02794 (ferritin)
 17,525 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chimeric immunotoxins that combine antigen recognition domains of antibodies and cytotoxic RNases have attracted much attention in recent years as potential targeted agents for cancer immunotherapy. In an attempt to obtain a structurally minimized immunofusion for folding/stability studies, we constructed the chimeric protein VL-barnase. The chimera comprises a small cytotoxic enzyme barnase, ribonuclease from Bacillus amyloliquefaciens, fused to the C-terminus of the light chain variable domain (VL) of the anti-human ferritin monoclonal antibody F11. While the individual VL domain was expressed in Escherichia coli as insoluble protein packed into inclusion bodies, its fusion to barnase resulted in a significant ( approximately 70%) fraction of soluble protein, with only a minor insoluble fraction ( approximately 30%) packed into inclusion bodies. The in vivo solubilizing effect of barnase was also observed in vitro and suggests a chaperone-like role that barnase exerted with regard to the N-terminal VL domain. Cytoplasmic VL-barnase was analyzed for structural and functional properties. The dimeric state of the chimeric protein was demonstrated by size-exclusion chromatography, thus indicating that fusion to barnase did not abrogate the intrinsic dimerization propensity of the VL domain. Ferritin-binding affinity and specificity in terms of constants of association with isoferritins were identical for the isolated VL domain and its barnase fusion, and RNase activity remained unchanged after the fusion. Intrinsic fluorescence spectra showed a fully compact tertiary structure of the fusion protein. However, significantly altered pH stability of the fusion protein versus individual VL and barnase was shown by the pH-induced changes in both intrinsic fluorescence and binding of ANS. Together, the results indicate that VL-barnase retained the antigen-binding affinity, specificity and RNase activity pertinent to the two individual constituents, and that their fusion into a single-chain chimeric protein resulted in an altered tertiary fold and pH stability.
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PMID:Fusion of the antiferritin antibody VL domain to barnase results in enhanced solubility and altered pH stability. 1498 41

Self-assembled particles of genetically engineered human L subunit ferritin expressing a silver-binding peptide were used as nanocontainers for the synthesis of silver nanoparticles. The inner cavity of the self-assembled protein cage displays a dodecapeptide that is capable of reducing silver ions to metallic silver. This chimeric protein cage when incubated in the presence of silver nitrate exhibits the growth of a silver nanocrystal within its cavity. Our studies indicate that it is possible to design chimeric cages, using specific peptide templates, for the growth of other inorganic nanoparticles.
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PMID:Engineered protein cages for nanomaterial synthesis. 1547 82

A chimeric protein VL-barstar that comprises the VL domain of anti-human ferritin monoclonal antibody F11 and barstar, the naturally occurring inhibitor of bacterial RNase barnase, has been constructed for study of structure-function characteristics of chimeric immunoglobulin fused proteins. Such chimeric constructs may be potentially employed for development of bivalent/bispecific antibodies on the basis of the high affinity interaction between barstar and barnase (the association constant is about 10(14) M(-1)). We have developed a protocol for VL-barstar expression in E. coli and purification and refolding from inclusion bodies that yields a homogeneous and soluble form of this protein. Differential scanning calorimetry in combination with fluorescence and CD spectroscopy revealed that the VL-barstar formed well-resolved ordered secondary and compact tertiary structures. However, partial loss of tertiary interactions resulted in low stability of the recombinant protein and the lack of functional activity of the two constituent modules. These conformational features suggest that the protein might be referred to the class of native molten globules, which comprises partially unfolded conformations stabilized under physiological conditions. Since individually expressed VL domain and barstar retain completely folded conformation and stable spatial structure, the incomplete folding of the chimeric protein may be attributed to interaction between heterologous domains, which appears at the folding stage preceding formation of a system of tertiary interactions in both structural modules. The results provide evidence for non-native interactions between heterologous modules that may occur in chimeric proteins composed of taxonomically distinct fusion partners.
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PMID:Folding and stability of chimeric immunofusion VL-barstar. 1552 8

A chimeric protein, VH-barnase, was obtained by fusing the VH domain of anti-human ferritin monoclonal antibody F11 to barnase, a bacterial RNase from Bacillus amyloliquefaciens. After refolding from inclusion bodies, the fusion protein formed insoluble aggregates. Off-pathway aggregation was significantly reduced by adding either purified GroEL/GroES chaperones or arginine, with 10-12-fold increase in the yield of the soluble protein. The final protein conformation was identical by calorimetric criteria and CD and fluorescence spectroscopy to that obtained without additives, thus suggesting that VH-barnase structure does not depend on folding conditions. Folding of VH-barnase resulted in a single calorimetrically revealed folding unit, the so-called "calorimetric domain", with conformation consistent with a molten globule that possessed well-defined secondary structure and compact tertiary conformation with partial exposure of hydrophobic patches and low thermodynamic stability. The unique feature of VH-barnase is that, despite the partially unfolded conformation and coupling into a single "calorimetric domain", this immunofusion retained both the antigen-binding and RNase activities that belong to the two heterologous domains.
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PMID:Fusion of barnase to antiferritin antibody F11 VH domain results in a partially folded functionally active protein. 1964 73

Human ferritins have been extensively studied to be used as nanocarriers for diverse applications and could represent a convenient alternative for targeted delivery of anticancer drugs and imaging agents. However, the most relevant limitation to their applications is the need for highly acidic experimental conditions during the initial steps of particle/cargo assembly, a process that could affect both drug stability and the complete reassembly of the ferritin cage. To overcome this issue the unique assembly of Archaeoglobus fulgidus ferritin was genetically engineered by changing a surface exposed loop of 12 amino acids connecting B and C helices to mimic the sequence of the analogous human H-chain ferritin loop. This new chimeric protein was shown to maintain the unique, cation linked, association-dissociation properties of Archaeoglobus fulgidus ferritin occurring at neutral pH values, while exhibiting the typical human H-homopolymer recognition by the transferrin receptor TfR1. The chimeric protein was confirmed to be actively and specifically internalized by HeLa cells, thus representing a unique nanotechnological tool for cell-targeted delivery of possible payloads for diagnostic or therapeutic purposes. Moreover, it was demonstrated that the 12 amino acids' loop is necessary and sufficient for binding to the transferrin receptor. The three-dimensional structure of the humanized Archaeoglobus ferritin has been obtained both as crystals by X-ray diffraction and in solution by cryo-EM.
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PMID:Humanized archaeal ferritin as a tool for cell targeted delivery. 2794 79

Hepcidin is a liver-synthesized hormone that plays a central role in the regulation of systemic iron homeostasis. To produce a new tool for its functional properties the cDNA coding for camel hepcidin-25 was cloned at the 5'end of human FTH sequence into the pASK-IBA43plus vector for expression in Escherichia coli The recombinant fusion hepcidin-ferritin-H subunit was isolated as an insoluble iron-containing protein. When alone it did not refold in a 24-mer ferritin molecule, but it did when renatured together with H- or L-ferritin chains. We obtained stable ferritin shells exposing about 4 hepcidin peptides per 24-mer shell. The molecules were then reduced and re-oxidized in a controlled manner to allow the formation of the proper hepcidin disulfide bridges. The functionality of the exposed hepcidin was confirmed by its ability to specifically bind the mouse macrophage cell line J774 that express ferroportin and to promote ferroportin degradation. This chimeric protein may be useful for studying the hepcidin-ferroportin interaction in cells and also as drug-delivery agent.
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PMID:Production and characterization of functional recombinant hybrid heteropolymers of camel hepcidin and human ferritin H and L chains. 2798 Jan 20

In this work, we have exploited the unique properties of a chimeric archaeal-human ferritin to encapsulate, deliver and release cytochrome c and induce apoptosis in a myeloid leukemia cell line. The chimeric protein combines the versatility in 24-meric assembly and cargo incorporation capability of Archaeglobus fulgidus ferritin with specific binding of human H ferritin to CD71, the "heavy duty" carrier responsible for transferrin-iron uptake. Delivery of ferritin-encapsulated cytochrome C to the Acute Promyelocytic Leukemia (APL) NB4 cell line, highly resistant to transfection by conventional methods, was successfully achieved in vitro. The effective liberation of cytochrome C within the cytosolic environment, demonstrated by double fluorescent labelling, induced apoptosis in the cancer cells.
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PMID:Ferritin nanovehicle for targeted delivery of cytochrome C to cancer cells. 3140 39