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Protein tyrosine phosphatase r (PTPr) belongs to the classical receptor type IIB family of protein tyrosine phosphatase, the most frequently mutated tyrosine phosphatase in human cancer. There are evidences to suggest that PTPr may act as a tumor suppressor gene and dysregulation of Tyr phosphorylation can be observed in diverse diseases, such as diabetes, immune deficiencies and cancer. PTPr variants in the catalytic domain have been identified in cancer tissues. These natural variants are nonsynonymous single nucleotide polymorphisms, variations of a single nucleotide occurring in the coding region and leading to amino acid substitutions. In this study we investigated the effect of amino acid substitution on the structural stability and on the activity of the membrane-proximal catalytic domain of PTPr. We expressed and purified as soluble recombinant proteins some of the mutants of the membrane-proximal catalytic domain of PTPr identified in colorectal cancer and in the single nucleotide polymorphisms database. The mutants show a decreased thermal and thermodynamic stability and decreased activation energy relative to phosphatase activity, when compared to wild- type. All the variants show three-state equilibrium unfolding transitions similar to that of the wild- type, with the accumulation of a folding intermediate populated at ∼4.0 M urea. [ABSTRACT FROM AUTHOR]

The stability of the dodecameric Listeria monocytogenes Dps has been compared with that of the Listeria innocua protein. The two proteins differ only in two amino acid residues that form an intersubunit salt-bridge in L. innocua Dps. This salt-bridge is replaced by a hydrogen bonding network in L. monocytogenes Dps as revealed by the X-ray crystal structure. The resistance to low pH and high temperature was assayed for both Dps proteins under equilibrium conditions and kinetically. Despite the identical equilibrium behavior, significant differences in the kinetic stability and activation energy of the unfolding process are apparent at pH 1.5. The higher stability of L. monocytogenes Dps has been accounted for in terms of the persistence of the hydrogen bonding network at this low pH value. In contrast, the salt-bridge between Lys 114 and Asp 126 characteristic of L. innocua Dps is most likely abolished due to protonation of Asp 126. Proteins 2007. © 2006 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Ferritin from the spleen of the Antarctic teleost Trematomus bernacchii is composed of a single subunit that contains both the ferroxidase center residues, typical of mammalian H chains, and the carboxylate residues forming the micelle nucleation site, typical of mammalian L chains. Comparison of the amino-acid sequence with those available from lower vertebrates indicates that T. bernacchii ferritin can be classified as an M-type homopolymer. Interestingly, the T. bernacchii ferritin chain shows 85.7% identity with a cold-inducible ferritin chain of the rainbow trout Salmo gairdneri . The structural and functional properties indicate that cold acclimation and functional adaptation to low temperatures are achieved without significant modification of the protein stability. In fact, the stability of T. bernacchii ferritin to denaturation induced by acid or temperature closely resembles that of mesophilic mammalian ferritins. Moreover iron is taken up efficiently and the activation energy of the reaction is 74.9 kJ·mol-1 , a value slightly lower than that measured for the human recombinant H ferritin (80.8 kJ·mol-1 ). [ABSTRACT FROM AUTHOR]

Investigates the stability of the dodecameric ferritin from gram positive bacteria, Listeria innocua. Stability of the ferritin tetracosamer at pH 2.0; Effect of anions on the protein at pH 2.0; Weakening of the quarternary structure at pH 1.5; Accessibility of hydrophobic residues.

Ferritin is characterized by a highly conserved architecture that comprises 24 subunits assembled into a spherical cage with 432 symmetry. The only known exception is the dodecameric ferritin from Listeria innocua. The structure of Listeria ferritin has been determined to a resolution of 2.35 Å by molecular replacement, using as a search model the structure of Dps from Escherichia coli. The Listeria 12-mer is endowed with 23 symmetry and displays the functionally relevant structural features of the ferritin 24-mer, namely the negatively charged channels along the three-fold symmetry axes that serve for iron entry into the cavity and a negatively charged internal cavity for iron deposition. The electron density map shows 12 iron ions on the inner surface of the hollow core, at the interface between monomers related by two-fold axes. Analysis of the nature and stereochemistry of the iron-binding ligands reveals strong similarities with known ferroxidase sites. The L. innocua ferritin site, however, is the first described so far that has ligands belonging to two different subunits and is not contained within a four-helix bundle. [ABSTRACT FROM AUTHOR]

Iron uptake and micelle formation in ferritin and apoferritin have been followed both spectrophotometrically and by means of sedimentation velocity experiments. Information was thus obtained on the molecular weight distribution of the reconstitution product. To achieve incorporation 'native' ferritin (whole ferritin as purified from horse spleen), 'native' apoferritin (apoferritin prepared by fractionation of ferritin preparations) and 'reduced' apoferritin (apoferritin prepared by reduction of ferritin by dithionite or ascorbic acid) have been incubated with ferrous salts in the presence of oxidizing agents under different experimental conditions. Although some iron is incorporated in 'native' ferritin, full saturation is not achieved and the molecular weight distribution of the incubated products remains heterogeneous. 'Native' and 'reduced' apoferritin show a similar iron incorporation, but the reconstitution products markedly differ in terms of their iron distribution. Ferritin reconstituted from 'native' apoferritin has a broad molecular weight distribution, while that reconstituted from 'reduced' apoferritin is characterized by a narrow, homogeneous molecular weight distribution. However treatment of apoferritin with reducing or oxidizing agents prior to the incubation alters the characteristics of the iron distribution without changing the iron incorporation properties. These results point to a role of the protein moiety not only in iron oxidation, but also in micelle formation. [ABSTRACT FROM AUTHOR]