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1.
The prion protein has garnered considerable interest because of its involvement in prion disease as well as its unresolved cellular function. The octarepeat region in the flexible N-domain is capable of binding copper through multiple coordination modes. Under conditions of low pH and low Cu2+ concentration, the four octarepeats (ORs) cooperatively coordinate a single copper ion. Based on the average structure of the PHGG and GWGQ portions of a copper-free OR2 model from molecular dynamics simulations, the starting structures of the OR4 complex could be constructed by assembling the repeating structure of PHGG and GWGQ fragments. The resulting model contains a preformed site suitable for Cu2+ coordination. Molecular dynamics simulations of Cu2+ bound to the assembled OR4 model (Cu:OR4) reveal a close association of specific Trp and Gly residues with the Cu2+ center. This low Cu2+-occupancy form of prion protein is redox-active and can readily initiate cleavage of the OR region, mediated by reactive oxygen species generated by Cu+. The OR region is known to be required for β-cleavage, as are the Trp residues within the OR region. The β-cleaved form of the prion protein accumulates in amyloid fibrils. Hence, the close approach of Trp and Gly residues to the Cu2+ coordination site in the low Cu2+-occupancy form of the OR region may signal an important interaction for the initiation of prion disease. 相似文献
2.
The native prion protein (PrP) has a two domain structure, with a globular folded α-helical C-terminal domain and a flexible
extended N-terminal region. The latter can selectively bind Cu2+ via four His residues in the octarepeat (OR) region, as well as two sites (His96 and His111) outside this region. In the
disease state, the folded C-terminal domain of PrP undergoes a conformational change, forming amorphous aggregates high in
β-sheet content. Cu2+ bound to the ORs can be redox active and has been shown to induce cleavage within the OR region, a process requiring conserved
Trp residues. Using computational modeling, we have observed that electron transfer from Trp residues to copper can be favorable.
These models also reveal that an indole-based radical cation or Cu+ can initiate reactions leading to protein backbone cleavage. We have also demonstrated, by molecular dynamics simulations,
that Cu2+ binding to the His96 and His111 residues in the remaining PrP N-terminal fragment can induce localized β-sheet structure,
allowing us to suggest a potential mechanism for the initiation of β-sheet misfolding in the C-terminal domain by Cu2+.
相似文献
Hans J. VogelEmail: |
3.
Sara Furlan Giovanni La Penna Francesco Guerrieri Silvia Morante Gian Carlo Rossi 《Journal of biological inorganic chemistry》2007,12(4):571-583
The human prion protein binds Cu2+ ions in the octarepeat domain of the N-terminal tail up to full occupancy at pH 7.4. Recent experiments have shown that the
HGGG octarepeat subdomain is responsible for holding the metal bound in a square-planar configuration. By using first principle
ab initio molecular dynamics simulations of the Car–Parrinello type, the coordination of copper to the binding sites of the
prion protein octarepeat region is investigated. Simulations are carried out for a number of structured binding sites. Results
for the complexes Cu(HGGGW)(wat), Cu(HGGG), and [Cu(HGGG)]2 are presented. While the presence of a Trp residue and a water molecule does not seem to affect the nature of the copper
coordination, high stability of the bond between copper and the amide nitrogen of deprotonated Gly residues is confirmed in
all cases. For the more interesting [Cu(HGGG)]2 complex, a dynamically entangled arrangement of the two domains with exchange of amide nitrogen bonds between the two copper
centers emerges, which is consistent with the short Cu–Cu distance observed in experiments at full copper occupancy. 相似文献
4.
del Pino P Weiss A Bertsch U Renner C Mentler M Grantner K Fiorino F Meyer-Klaucke W Moroder L Kretzschmar HA Parak FG 《European biophysics journal : EBJ》2007,36(3):239-252
The cellular prion protein (PrPC) is a Cu2+ binding protein connected to the outer cell membrane. The molecular features of the Cu2+ binding sites have been investigated and characterized by spectroscopic experiments on PrPC-derived peptides and the recombinant human full-length PrPC (hPrP-[23-231]). The hPrP-[23-231] was loaded with 63Cu under slightly acidic (pH 6.0) or neutral conditions. The PrPC/Cu2+-complexes were investigated by extended X-ray absorption fine structure (EXAFS), electron paramagnetic resonance (EPR), and
electron nuclear double resonance (ENDOR). For comparison, peptides from the copper-binding octarepeat domain were investigated
in different environments. Molecular mechanics computations were used to select sterically possible peptide/Cu2+ structures. The simulated EPR, ENDOR, and EXAFS spectra of these structures were compared with our experimental data. For
a stoichiometry of two octarepeats per copper the resulting model has a square planar four nitrogen Cu2+ coordination. Two nitrogens belong to imidazole rings of histidine residues. Further ligands are two deprotonated backbone
amide nitrogens of the adjacent glycine residues and an axial oxygen of a water molecule. Our complex model differs significantly
from those previously obtained for shorter peptides. Sequence context, buffer conditions and stoichiometry of copper show
marked influence on the configuration of copper binding to PrPC.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
5.
We studied the redox behavior of copper-binding sites in prion protein (PrP) to clarify copper’s role in the pathological
mechanism underlying prion diseases. We investigated the coordination structures, binding affinities, and redox potentials
of copper-binding peptide fragments derived from the N-terminal domain of PrP by density functional theory calculations. We
used four models for copper-binding moieties in PrP(60–96): two were derived from the PHGGGWGQ octapeptide repeat region of
PrP(60–91), and the others were tripeptide Gly-Thr-His fragments derived from the copper-binding nonoctarepeat site around
His96. We found that such PrP-derived copper-binding complexes exhibit conformationally dependent redox behavior; for example,
the copper-binding complex derived from the octarepeat region tends to possess high reduction potential for the Cu(II)/Cu(I)
couple, exceeding 0 V versus the standard hydrogen electrode, whereas the copper-binding nonoctarepeat model around His96
tends to possess high oxidation potential for the Cu(II)/Cu(III) couple and stabilize the higher-valent Cu(III) state. It
is possible that such distinct redox activities of a copper-binding PrP are involved in the mechanism underlying prion diseases. 相似文献
6.
Jones CE Klewpatinond M Abdelraheim SR Brown DR Viles JH 《Journal of molecular biology》2005,346(5):1393-1407
The prion protein (PrP) is a Cu2+ binding cell surface glyco-protein. Misfolding of PrP into a beta-sheet rich conformation is associated with transmissible spongiform encephalopathies. Here we use Ni2+ as a diamagnetic probe to further understand Cu2+ binding to PrP. Like Cu2+, Ni2+ preferentially binds to an unstructured region between residues 90 and 126 of PrP, which is a key region for amyloidogenicity and prion propagation. Using both 1H NMR and visible-circular dichroism (CD) spectroscopy, we show that two Ni2+ ions bind to His96 and His111 independently of each other. 1H NMR indicates that both Ni2+ binding sites form square-planar diamagnetic complexes. We have previously shown that Cu2+ forms a paramagnetic square-planar complex in this region, suggesting that Ni2+ could be used as a probe for Cu2+ binding. In addition, competition studies show that two Cu2+ ions can displace Ni2+ from these sites. Upon Ni2+ addition 1H NMR changes in chemical shifts indicate the imidazole ring and amide nitrogen atoms to the N terminus of both His96 and His111 act as coordinating ligands. Use of peptide fragments confirm that PrP(92-96) and PrP(107-111) represent the minimal binding motif for the two Ni2+ binding sites. Analysis of Cu2+ loaded visible-CD spectra show that as with Ni2+, PrP(90-115) binds two Cu2+ ions at His96 and His111 independently of each other. Visible CD studies with PrP(23-231Delta51-90), a construct of PrP(23-231) with the octarepeat region deleted to improve solubility, confirm binding of Ni2+ to His96 and His111 in octarepeat deleted PrP(23-231). The structure of the Cu/Ni complexes is discussed in terms of the implications for prion protein function and disease. 相似文献
7.
Copper(II)-induced secondary structure changes and reduced folding stability of the prion protein 总被引:1,自引:0,他引:1
Younan ND Klewpatinond M Davies P Ruban AV Brown DR Viles JH 《Journal of molecular biology》2011,410(3):369-382
The cellular isoform of the prion protein PrPC is a Cu2+-binding cell surface glycoprotein that, when misfolded, is responsible for a range of transmissible spongiform encephalopathies. As changes in PrPC conformation are intimately linked with disease pathogenesis, the effect of Cu2+ ions on the structure and stability of the protein has been investigated. Urea unfolding studies indicate that Cu2+ ions destabilise the native fold of PrPC. The midpoint of the unfolding transition is reduced by 0.73 ± 0.07 M urea in the presence of 1 mol equiv of Cu2+. This equates to an appreciable difference in free energy of unfolding (2.02 ± 0.05 kJ mol− 1 at the midpoint of unfolding). We relate Cu2+-induced changes in secondary structure for full-length PrP(23-231) to smaller Cu2+ binding fragments. In particular, Cu2+-induced structural changes can directly be attributed to Cu2+ binding to the octarepeat region of PrPC. Furthermore, a β-sheet-like transition that is observed when Cu ions are bound to the amyloidogenic fragment of PrP (residues 90-126) is due only to local Cu2+ coordination to the individual binding sites centred at His95 and His110. Cu2+ binding does not directly generate a β-sheet conformation within PrPC; however, Cu2+ ions do destabilise the native fold of PrPC and may make the transition to a misfolded state more favourable. 相似文献
8.
The prion protein is a ubiquitous neuronal membrane protein. Misfolding of the prion protein has been implicated in transmissible spongiform encephalopathies (prion diseases). It has been demonstrated that the human prion protein (PrP) is capable of coordinating at least five CuII ions under physiological conditions; four copper binding sites can be found in the octarepeat domain between residues 61 and 91, while another copper binding site can be found in the unstructured “amyloidogenic” domain between residues 91 and 126 PrP(91-126). Herein we expand upon a previous study [J. Shearer, P. Soh, Inorg. Chem. 46 (2007) 710-719] where we demonstrated that the physiologically relevant high affinity CuII coordination site within PrP(91-126) is found between residues 106 and 114. It was shown that CuII is contained within a square planar (N/O)3S coordination environment with one His imidazole ligand (H(111)) and one Met thioether ligand (either M(109) or M(112)). The identity of the Met thioether ligand was not identified in that study. In this study we perform a detailed investigation of the CuII coordination environment within the PrP fragment containing residues 106-114 (PrP(106-114)) involving optical, X-ray absorption, EPR, and fluorescence spectroscopies in conjunction with electronic structure calculations. By using derivatives of PrP(106-114) with systematic Met → Ile “mutations” we show that the CuII coordination environment within PrP(106-114) is actually comprised of a mixture of two major species; one CuII(N/O)3S center with the M(109) thioether coordinated to CuII and another CuII(N/O)3S center with the M(112) thioether coordinated to CuII. Furthermore, deletion of one or more Met residues from the primary sequence of PrP(106-114) both reduces the CuII affinity of the peptide by two to seven fold, and renders the resulting CuII metallopeptides redox inactive. The biological implications of these findings are discussed. 相似文献
9.
Carina Treiber Andrew R. Thompsett Rüdiger Pipkorn David R. Brown Gerd Multhaup 《Journal of biological inorganic chemistry》2007,12(5):711-720
The prion protein (PrP) is a metalloprotein with an unstructured region covering residues 60–91 that bind two to six Cu(II)
ions cooperatively. Cu can bind to PrP regions C-terminally to the octarepeat region involving residues His111 and/or His96.
In addition to Cu(II), PrP binds Zn(II), Mn(II) and Ni(II) with binding constants several orders of magnitudes lower than
those determined for Cu. We used for the first time surface plasmon resonance (SPR) analysis to dissect metal binding to specific
sites of PrP domains and to determine binding kinetics in real time. A biosensor assay was established to measure the binding
of PrP-derived synthetic peptides and recombinant PrP to nitrilotriacetic acid chelated divalent metal ions. We have identified
two separate binding regions for binding of Cu to PrP by SPR, one in the octarepeat region and the second provided by His96
and His111, of which His96 is more essential for Cu coordination. The octarepeat region at the N-terminus of PrP increases
the affinity for Cu of the full-length protein by a factor of 2, indicating a cooperative effect. Since none of the synthetic
peptides covering the octarepeat region bound to Mn and recombinant PrP lacking this sequence were able to bind Mn, we propose
a conformational binding site for Mn involving residues 91–230. A novel low-affinity binding site for Co(II) was discovered
between PrP residues 104 and 114, with residue His111 being the key amino acid for coordinating Co(II). His111 is essential
for Co(II) binding, whereas His96 is more important than His111 for binding of Cu(II). 相似文献
10.
Jones CE Abdelraheim SR Brown DR Viles JH 《The Journal of biological chemistry》2004,279(31):32018-32027
The prion protein (PrP) is a Cu(2+) binding cell surface glycoprotein that can misfold into a beta-sheet-rich conformation to cause prion diseases. The majority of copper binding studies have concentrated on the octarepeat region of PrP. However, using a range of spectroscopic techniques, we show that copper binds preferentially to an unstructured region of PrP between residues 90 and 115, outside of the octarepeat domain. Comparison of recombinant PrP with PrP-(91-115) indicates that this prion fragment is a good model for Cu(2+) binding to the full-length protein. In contrast to previous reports we show that Cu(2+) binds to this region of PrP with a nanomolar dissociation constant. NMR and EPR spectroscopy indicate a square-planar or square-pyramidal Cu(2+) coordination utilizing histidine residues. Studies with PrP analogues show that the high affinity site requires both His(96) and His(111) as Cu(2+) ligands, rather than a complex centered on His(96) as has been previously suggested. Our circular dichroism studies indicate a loss of irregular structure on copper coordination with an increase in beta-sheet conformation. It has been shown that this unstructured region, between residues 90 and 120, is vital for prion propagation and different strains of prion disease have been linked with copper binding. The role of Cu(2+) in prion misfolding and disease must now be re-evaluated in the light of these findings. 相似文献
11.
Francesco Guerrieri Velia Minicozzi Silvia Morante Giancarlo Rossi Sara Furlan Giovanni La Penna 《Journal of biological inorganic chemistry》2009,14(3):361-374
The octarepeat region of the prion protein can bind Cu2+ ions up to full occupancy (one ion per octarepeat) at neutral pH. While crystallographic data show that the HGGG octarepeat
subdomain is the basic binding unit, multiple histidine coordination at lower Cu occupancy has been reported by X-ray absorption
spectroscopy, EPR, and potentiometric experiments. In this paper we investigate, with first principles Car–Parrinello simulations,
the first step for the formation of the Cu low-level binding mode, where four histidine side chains are coordinated to the
same Cu2+ ion. This step involves the further binding of a second histidine to an already HGGG domain bonded Cu2+ ion. The influence of the pH on the ability of Cu to bind two histidine side chains was taken into account by simulating
different protonation states of the amide N atoms of the two glycines lying nearest to the first histidine. Multiple histidine
coordination is also seen to occur when glycine deprotonation occurs and the presence of the extra histidine stabilizes the
Cu–peptide complex. Though the stabilization effect slightly decreases with the number of deprotonated glycines (reaching
a minimum when both N atoms of the two nearest glycines are available as Cu ligands), the system is still capable of binding
the second histidine in a 4N tetrahedral (though slightly distorted) coordination, whose energy is very near to that of the
crystallographic square-planar 3N1O coordination. This result suggests that at low metal concentration the reorganization
energy associated with Cu(II)/Cu(I) reduction is small also at pH ~ 7, when glycines are deprotonated.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.
相似文献
Giovanni La PennaEmail: |
12.
Ildikó Turi Dorina Szikszai Giuseppe Di Natale Enrico Rizzarelli Imre Sóvágó 《Journal of inorganic biochemistry》2010,104(8):885-4348
Nickel(II) complexes of the peptide fragments of human prion protein containing histidyl residues both inside and outside the octarepeat domain have been studied by the combined application of potentiometric, UV-visible and circular dichroism spectroscopic methods. The imidazole-N donor atoms of histidyl residues are the exclusive metal binding sites below pH 7.5, but the formation of stable macrochelates was characteristic only for the peptide HuPrP(76-114) containing four histidyl residues. Yellow colored square planar complexes were obtained above pH 7.5-8 with the cooperative deprotonation of three amide nitrogens in the [Nim,N−,N−,N−] coordination mode. It was found that the peptides can bind as many nickel(II) ions as the number of independent histidyl residues. All data supported that the complex formation processes of nickel(II) are very similar to those of copper(II), but with a significantly reduced stability for nickel(II), which shifts the complex formation reactions into the slightly alkaline pH range. The formation of coordination isomers was characteristic of the mononuclear complexes with a significant preference for the nickel(II) binding at the histidyl sites outside the octarepeat domain. The results obtained for the two-histidine fragments of the protein, HuPrP(91-115), HuPrP(76-114)H85A and HuPrP(84-114)H96A, made it possible to compare the binding ability of the His96 and His111 sites. These data reveal a significant difference in the nickel(II) and copper(II) binding sites of the peptides: His96 was found to predominate almost completely for nickel(II) ions, while the opposite order, but with comparable concentrations, was reported for copper(II). 相似文献
13.
CopH is a periplasmic copper-binding protein from Cupriavidus metallidurans CH34 that contains two histidine residues. Both His24 and His26 contribute to the formation of two high-affinity copper-binding sites in wild-type CopH and are likely involved in a 2N2O coordination sphere in the equatorial plane. We have used site-directed mutagenesis, and a series of spectroscopic and calorimetric studies to further characterize the copper-binding sites in CopH. While His24 plays a predominant role in copper affinity, one Cu-binding site was lost when either histidine residue was mutated. However, as shown by NMR and EPR, the mutation of the His residues does not affect the structural organization of the Cu-binding site nor the number of nitrogen ligands involved in copper ligation. In the absence of structural data, we propose a model that conciliates most of the spectroscopic data recorded during this study. 相似文献
14.
Daniela Valensin Karolina Gajda Gianni Valensin Henryk Kozlowski 《Journal of inorganic biochemistry》2010,104(1):71-2450
Both human (h) and chicken (Ch) prion proteins (PrP) bind copper ions within the so called “tandem repeat” N-terminal region. Outside this region, hPrP possesses two additional copper binding sites, localized at His-96 and His-111 in the so called “amylodogenic” or neurotoxic region (residues 91-126). Also ChPrP possesses a similar region (ChPrP105−140) containing two His (His-110 and His-124) and an identical hydrophobic tail of 15 amino acids rich in Ala and Gly. The copper binding abilities within such region of ChPrP were investigated by NMR, CD and potentiometry using Ni2+ as diamagnetic probe. The formation of diamagnetic metal complexes allowed to monitor the chemical shift and signal intensity variations and to determine the structural and kinetic features of the His-110 and His-124 metal binding sites. Finally a comparison between the hPrP and ChPrP metal binding abilities was performed. We found that the two prion proteins exhibited different copper and nickel preferences with the favoured metal binding sites localized at opposite His: His-110 for ChPrP, and His-111 for hPrP. 相似文献
15.
James S. Hyde Brian Bennett Glenn L. Millhauser William E. Antholine 《Biophysical journal》2009,96(8):3354-3362
A double octarepeat prion protein construct, which has two histidines, mixed with copper sulfate in a 3:2 molar ratio provides at most three imidazole ligands to each copper ion to form a square-planar Cu2+ complex. This work is concerned with identification of the fourth ligand. A new (to our knowledge) electron paramagnetic resonance method based on analysis of the intense features of the electron paramagnetic resonance spectrum in the g⊥ region at 2 GHz is introduced to distinguish between three and four nitrogen ligands. The methodology was established by studies of a model system consisting of histidine imidazole ligation to Cu2+. In this spectral region at 2 GHz (S-band), g-strain and broadening from the possible rhombic character of the Zeeman interaction are small. The most intense line is identified with the MI = +1/2 extra absorption peak. Spectral simulation demonstrated that this peak is insensitive to cupric Ax and Ay hyperfine interaction. The spectral region to the high-field side of this peak is uncluttered and suitable for analysis of nitrogen superhyperfine couplings to determine the number of nitrogens. The spectral region to the low-field side of the intense extra absorption peak in the g⊥ part of the spectrum is sensitive to the rhombic distortion parameters Ax and Ay. Application of the method to the prion protein system indicates that two species are present and that the dominant species contains four nitrogen ligands. A new loop-gap microwave resonator is described that contains ∼1 mL of frozen sample. 相似文献
16.
Michael H. Suhre Simone Hess Adrian V. Golser 《Journal of inorganic biochemistry》2009,103(12):1711-1720
There is a large body of evidence that divalent metal ions, particularly copper, might play a role in several protein folding pathologies like Alzheimer’s disease, Parkinson’s disease or the prion diseases. However, contribution of metal ions on pathogenesis and their molecular influence on the formation of amyloid structures is not clear. Therefore, the general influence of metals on the formation of amyloids is still controversially discussed. We have utilized the well established system of yeast Sup35p-NM to investigate the role of three different metal ions, Cu2+, Mn2+ and Zn2+, on amyloidogenesis. Recently, it has been shown that the prion determining region NM of the Saccharomyces cerevisiae prion protein Sup35p, which is responsible for the yeast prion phenotype [PSI+], specifically binds Cu2+ ions. We further characterized the affinity of NM for Cu2+, which were found to be comparable to that of other amyloidogenic proteins like the mammalian prion protein PrP. The specific binding sites could be located in the aminoterminal N-region which is known to initiate formation of amyloidogenic nuclei. In the presence of Cu2+, fibril nucleation was significantly delayed, probably due to influences of copper on the oligomeric ensemble of soluble Sup35p-NM, since Cu2+ altered the tertiary structure of soluble Sup35p-NM, while no influences on fibril elongation could be detected. The secondary structure of soluble or fibrous protein and the morphology of the fibrils were apparently not altered when assembled in presence of Cu2+. In contrast, Mn2+ and Zn2+ did not bind to Sup35p-NM and did not exhibit significant effects on the formation of NM amyloid fibrils. 相似文献
17.
Cu2+ is an essential metal ion that plays a critical role in the regulation of a number of ion channels and receptors in addition to acting as a cofactor in a variety of enzymes. Here, we showed that human melastatin transient receptor potential 2 (hTRPM2) channel is sensitive to inhibition by extracellular Cu2+. Cu2+ at concentrations as low as 3 µM inhibited the hTRPM2 channel completely and irreversibly upon washing or using Cu2+ chelators, suggesting channel inactivation. The Cu2+-induced inactivation was similar when the channels conducted inward or outward currents, indicating the permeating ions had little effect on Cu2+-induced inactivation. Furthermore, Cu2+ had no effect on singe channel conductance. Alanine substitution by site-directed mutagenesis of His995 in the pore-forming region strongly attenuated Cu2+-induced channel inactivation, and mutation of several other pore residues to alanine altered the kinetics of channel inactivation by Cu2+. In addition, while introduction of the P1018L mutation is known to result in channel inactivation, exposure to Cu2+ accelerated the inactivation of this mutant channel. In contrast with the hTRPM2, the mouse TRPM2 (mTRPM2) channel, which contains glutamine at the position equivalent to His995, was insensitive to Cu2+. Replacement of His995 with glutamine in the hTRPM2 conferred loss of Cu2+-induced channel inactivation. Taken together, these results suggest that Cu2+ inactivates the hTRPM2 channel by interacting with the outer pore region. Our results also indicate that the amino acid residue difference in this region gives rise to species-dependent effect by Cu2+ on the human and mouse TRPM2 channels. 相似文献
18.
He Song Andrew C. Weitz Michael P. Hendrich Edwin A. Lewis Joseph P. Emerson 《Journal of biological inorganic chemistry》2013,18(6):595-598
Reengineering metalloproteins to generate new biologically relevant metal centers is an effective a way to test our understanding of the structural and mechanistic features that steer chemical transformations in biological systems. Here, we report thermodynamic data characterizing the formation of two type-2 copper sites in carbonic anhydrase and experimental evidence showing one of these new, copper centers has characteristics similar to a variety of well-characterized copper centers in synthetic models and enzymatic systems. Human carbonic anhydrase II is known to bind two Cu2+ ions; these binding events were explored using modern isothermal titration calorimetry techniques that have become a proven method to accurately measure metal-binding thermodynamic parameters. The two Cu2+-binding events have different affinities (K a approximately 5 × 1012 and 1 × 1010), and both are enthalpically driven processes. Reconstituting these Cu2+ sites under a range of conditions has allowed us to assign the Cu2+-binding event to the three-histidine, native, metal-binding site. Our initial efforts to characterize these Cu2+ sites have yielded data that show distinctive (and noncoupled) EPR signals associated with each copper-binding site and that this reconstituted enzyme can activate hydrogen peroxide to catalyze the oxidation of 2-aminophenol. 相似文献
19.
《Free radical biology & medicine》2009,46(12):1622-1630
The cellular prion protein (PrPc) plays a crucial role in the pathogenesis of prion diseases, but its physiological function is far from understood. Several candidate functions have been proposed including binding and internalization of metal ions, a superoxide dismutase-like activity, regulation of cellular antioxidant activities, and signal transduction. The transmembrane (TM1) region of PrPc (residues 110–135) is particularly interesting because of its very high evolutionary conservation. We investigated a possible role of TM1 in the antioxidant defense, by assessing the impact of overexpressing wt-PrP or deletion mutants in N2A mouse neuroblastoma cells on intracellular reactive oxygen species (ROS) levels. Under conditions of oxidative stress, intracellular ROS levels were significantly lowered in cells overexpressing either wild-type PrPc (wt-PrP) or a deletion mutant affecting TM1 (Δ8TM1-PrP), but, as expected, not in cultures overexpressing a deletion mutant lacking the octapeptide region (Δocta-PrP). Overexpression of wt-PrP, Δ8TM1-PrP, or Δocta-PrP did not affect basal ROS levels. Interestingly, the mitochondrial membrane potential was significantly lowered in Δocta-PrP-transfected cultures in the absence of oxidative stress. We conclude that the protective effect of PrPc against oxidative stress involves the octarepeat region but not the TM1 domain nor the high-affinity copper binding site described for human residues His96/His111. 相似文献
20.
We study via all atom classical molecular dynamics (MD) simulation the process of uptake of ferrous ions (Fe2+) into the human ferritin protein and the catalytic ferroxidase sites via pores (“channels”) in the interior of the protein. We observe that the three‐fold hydrophilic channels serve as the main entrance pathway for the Fe2+ ions. The binding sites along the ion pathway are investigated. Two strong binding sites, at the Asp131 and Glu134 residues and two weak binding sites, at the His118 and Cys130 are observed inside the three‐fold channel. We also identify an explicit pathway for an ion exiting the channel into the central core of the protein as it moves to the ferroxidase site. The diffusion of an Fe2+ ion from the inner opening of the channel to a ferroxidase site located in the interior region of the protein coat is assisted by Thr135, His136 and Tyr137. The Fe2+ ion binds preferentially to site A of the ferroxidase site. © 2013 Wiley Periodicals, Inc. 相似文献