1,721,025 research outputs found
Biological activity and pathological implications of misfolded proteins.
No full textThe physiological metabolism of proteins guarantees that different cellular compartments contain the appropriate concentration of proteins to perform their biological functions and, after a variable period of wear and tear, mediates their natural catabolism. The equilibrium between protein synthesis and catabolism ensures an effective turnover, but hereditary or acquired abnormalities of protein structure can provoke a premature loss of biological function, an accelerated catabolism and diseases caused by the loss of an irreplaceable function. In certain proteins, abnormal structure and metabolism are associated with a strong tendency to self-aggregation into a polymeric fibrillar structure, and in these cases the disease is not principally caused by the loss of an irreplaceable function but by the action of this new biological entity. Amyloid fibrils are an apparently inert, insoluble, mainly extracellular protein polymer that kills the cell without tissue necrosis but by activation of the apoptotic mechanism. We analyzed the data reported so far on the structural and functional properties of four prototypic proteins with well-known biological functions (lysozyme, transthyretin, beta 2-microglobulin and apolipoprotein AI) that are able to create amyloid fibrils under certain conditions, with the perspective of evaluating whether the achievement of biological function favors or inhibits the process of fibril formation. Furthermore, studying the biological functions carried out by amyloid fibrils reveals new types of protein-protein interactions in the transmission of messages to cells and may provide new ideas for effective therapeutic strategies
Immunoglobulin light chain amyloidosis--the archetype of structural and pathogenic variability.
No full textAL amyloidosis is caused by deposition in target tissue of amyloid fibrils constituted by monoclonal immunoglobulin light chains. The amyloidogenic plasma cells derive from a transformed memory B cell that can be identified by anti-idiotype monoclonal antibodies. Comparison of the primary structures of amyloidogenic and nonamyloidogenic light chains does not show any common structural motif in the amyloidogenic variants but reveals peculiar replacements which can destabilize the folding state. Reduced folding stability now appears to be a unifying property of amyloidogenic light chains. The tendency of these proteins to populate a partially unfolded intermediate state is a key event in the self-association that progresses to the formation of oligomers and fibrils. The mechanism of organ damage caused by AL amyloid deposition is not known, but clinical findings suggest that the process of amyloid fibril formation itself exerts tissue toxic effects independently of the amount of amyloid deposited. Since the disease is caused by the neoplastic expansion of the plasma cell population synthesizing the amyloidogenic light chains, the clone represents the prime therapeutic target of conventional chemotherapy and experimental immunotherapy. In common with other types of amyloidosis the therapeutic strategy can take advantage of drugs able to improve the reabsorption of the amyloid deposits or able to bind and stabilize the light chain in the native-like folded state
Use of anti-(beta2 microglobulin) mAb to study formation of amyloid fibrils
No full textThree mAbs, IgG1k 1F11, 7B6 and 14H3, were raised against in vitro-self-aggregated beta2-microglobulin. They recognize the native and unfolded forms of the protein and bind its fibrillar form that is present in amyloid tissue. When assayed in fibrillogenesis tests in vitro, mAb 14H3 inhibited fibril formation from beta2-microglobulin. This mAb recognizes a sequential epitope corresponding to the C-terminal octapeptide, residues 92-99, of beta2-microglobulin. By using synthetic peptides it has been found that the integrity of the sequence is essential for the formation of the immunocomplex: the binding affinity is lowered by one order of magnitude (Kd from 10(-7) M to 10(-6) M) by removal of Met99 and completely abolished when both Asp98 and Met99 are lost or Arg98 is substituted with Lys. The other two mAbs, 1F11 and 7B6, which bind sequences 20-41 and 63-75, respectively, are without effect on beta2-microglobulin fibrillogenesis. These two mAbs recognize beta2-microglobulin bound to the heavy chain in the major histocompatibility complex of type I located in the cell membrane, a property which is not shared by mAb 14H3
Preliminary crystallographic characterization of the human beta2 microglobulin His31Tyr mutant in a tetrameric assembly.
No full textPatients receiving prolonged haemodialysis treatment are exposed to a variety of arthropathies and bone lesions arising from deposition of amyloid material in the skeletal system. beta2 microglobulin is the 11.7 kDa light chain of the class I major histocompatibility complex, from which it is normally released to plasmatic fluids, transported to kidneys and excreted. Owing to renal failure it accumulates, giving rise to dialysis-related amyloidosis, a severe disease found in patients receiving dialysis for several years. The three-dimensional structure of beta2 microglobulin is known to be based on a seven-stranded beta-sandwich fold, typical of the class C immunoglobulin superfamily. Analysis of the protein fold in different mutants and/or crystal environments and of its structural stability may help in understanding the molecular bases of amyloid fibril formation and of diseases related to protein misfolding. Here, the preliminary crystallographic analysis of the His31Tyr beta2 microglobulin mutant, designed to abolish the copper-ion binding observed in the wild-type protein, is presented. The protein mutant displays increased fold stability, faster folding kinetics and crystallizes in the tetragonal C222(1) space group, with unit-cell parameters a = 105.2, b = 150.2, c = 93.7 A and four molecules per asymmetric unit
Structural and functional characterization of three human immunoglobulin κ light chains with different pathological implications
No full textAbstractThe structural properties of three immunoglobulins light chains: κ SCI, responsible for light chain deposition disease (Bellotti, V., Stoppini, M., Merlini, G., Zapponi, M.C., Meloni, M.L., Banfi, G. and Ferri, G. (1991) Biochim. Biophys. Acta 1097, 177–182), k INC responsible for light chain amyloidosis (Ferri, G., Stoppini, M., Iadarola, P., Bellotti, V. and Merlini, G. (1989) Biochim. Biophys. Acta 995, 103–108) and the non-pathogenic κ MOS were analyzed by fluorescence spectroscopy and circular dichroism.Comparative evaluation of the data shows that SCI and MOS have similar stability under different conditions, while the amyloid k INC behaves as a very unstable protein.As calculated from the GdnHCl curves, the midpoint of unfolding transition was 1.35 M for SCI, 1.20 M for MOS and 0.1 M for INC. Analysis of CD spectra evidences that the three proteins conserve their conformation in the range of pH 4–8. Change in temperature at pH 4.0 produces the premature transition of INC (Tm 40°C) with respect to SCI and MOS (Tm 50°C). At this pH both the pathological SCI and INC light chains aggregate at a temperature of 20°C lower than the normal counterpart. The specific kidney deposition of κ SCI has been evidenced after injection of the 125I labelled light chain into mice. No deposition was detectable in the case of INC and MOS
DETECTION OF FRAGMENTS OF BETA2-MICROGLOBULIN IN AMYLOID FIBRILS
No full textA major controversy regarding dialysis related amyloidosis (DRA) is about the presence and the role of truncated forms of beta2-microglobulin (beta2M) in the amyloid fibrils. Whereas Linke et al have reported the presence of proteolyzed forms of beta2M in the amyloid deposits in over 12 cases, other groups have not confirmed this finding. A similar analysis conduced in our laboratory with beta2-fibrils obtained from six patients confirms Linke's data
Cinetiche di refolding e studi di affinità mediante elettroforesi capillare: effetto di ligandi di interesse farmaceutico sulla conformazione della Beta 2-microglobulina
No full textProduction and immunicytochemical characterization of monoclonal antibodies to SA gene product
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