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Jim Peterson

Publications and source records attributed to Jim Peterson.

13 recordsLinked to original sources
PubMed2006

Peroxidase activity and structural transitions of cytochrome c bound to cardiolipin-containing membranes.

During apoptosis, cytochrome c (cyt c) is released from intermembrane space of mitochondria into the cytosol where it triggers the caspase-dependent machinery. We discovered that cyt c plays another critical role in early apoptosis as a cardiolipin (CL)-specific oxygenase to produce CL hydroperoxides required for release of pro-apoptotic factors [Kagan, V. E., et al. (2005) Nat. Chem. Biol. 1, 223-232]. We quantitatively characterized the activation of peroxidase activity of cyt c by CL and hydrogen peroxide. At low ionic strength and high CL/cyt c ratios, peroxidase activity of the CL/cyt c complex was increased >50 times. This catalytic activity correlated with partial unfolding of cyt c monitored by Trp(59) fluorescence and absorbance at 695 nm (Fe-S(Met(80)) band). The peroxidase activity increase preceded the loss of protein tertiary structure. Monounsaturated tetraoleoyl-CL (TOCL) induced peroxidase activity and unfolding of cyt c more effectively than saturated tetramyristoyl-CL (TMCL). TOCL/cyt c complex was found more resistant to dissociation by high salt concentration. These findings suggest that electrostatic CL/cyt c interactions are central to the initiation of the peroxidase activity, while hydrophobic interactions are involved when cyt c's tertiary structure is lost. In the presence of CL, cyt c peroxidase activity is activated at lower H(2)O(2) concentrations than for isolated cyt c molecules. This suggests that redistribution of CL in the mitochondrial membranes combined with increased production of H(2)O(2) can switch on the peroxidase activity of cyt c and CL oxidation in mitochondria-a required step in execution of apoptosis.

Acridine Orange↗
PubMed2005

Nitrosative stress results in irreversible inhibition of purified mitochondrial complexes I and III without modification of cofactors.

The effects of both nitric oxide (NO) and peroxynitrite on complexes I (NADH dehydrogenase) and III (cytochrome c reductase) isolated from bovine heart have been examined. EPR signals ("g=2.01") previously detected in association with loss of complex I and III activities in cultured cells and isolated mitochondria subjected to nitrosative stress are shown not to arise from these particular enzymes. Neither NO nor peroxynitrite (ONO(2)(-)) reacts to any appreciable extent with the oxidized forms of flavin mononucleotide, iron-sulfur clusters, or heme moieties found in complexes I and III. However, ONO(2)(-) is readily able to abstract electrons from the reduced forms of both complexes I and III, without any apparent modification of the enzyme cofactors. While no attempt was made in the present study to catalog all the possible modifications, it is clear that ONO(2)(-) can react with the protein moieties of the enzymes. For example, when added in excess, ONO(2)(-) derivatizes a select few tyrosine residues in both complexes I and III forming 3-nitrotyrosine as detected by immunoblots. In the case of complex I, we find a minimum of 3 out of the 46 subunits present were modified (49, approximately 18, and approximately 15kDa); whereas in complex III, 4 out of the 13 subunits stained for 3-nitrotyrosine (46, 27, 7, and 6kDa). Significant irreversible inhibition of activity required the addition of >10(2)-fold excesses of ONO(2)(-) to the enzymes. At 10(3)-fold excess of added ONO(2)(-), the activity of complex I was only diminished by approximately 18%, while a 60% loss of activity was observed for complex III.

Animals↗
PubMed2005

Glutathione depletion renders rat hepatocytes sensitive to nitric oxide donor-mediated toxicity.

Nitric oxide (NO) can be either cytoprotective or cytotoxic in hepatocytes, depending on conditions within the cell. We hypothesized that redox status is a determinant of NO effects on cell viability. To cause the disturbance of redox homeostasis in the hepatocytes, cells were treated with the following glutathione (GSH) depleting agents: (1) chronic depletion by 18 hours pretreatment with buthionine sulfoximine (BSO), which depletes GSH by blocking its biosynthesis; and (2) acute depletion by 1 hour pretreatment with diethyl maleate (DEM), which conjugates GSH by the GSH-S-transferase catalyzed reaction. S-nitroso-N-acetyl-D,L-penicillamine (SNAP), a NO donor, was added after removal of GSH-depleting agents. Individual treatment with either SNAP or GSH depletion did not appreciably affect viability. A significant increase of cytotoxicity in hepatocytes was observed with the combination of a concentration and time course regimen of SNAP and GSH depletion. SNAP treatment of GSH-depleted hepatocytes led to an increase in LDH release and oxidative stress, disruption of mitochondrial membrane potential, the presence of nitrotyrosine (an indicator of peroxynitrite (ONOO-) generation), and a decrease in adenosine triphosphate (ATP) content. The interference of mitochondrial respiratory enzymes, especially with the combination treatments, indicated different levels of disturbance of electron transfer, superoxide generation, and ATP production. Other commonly used NO donors were found to exhibit lower and slower toxicity in the setting of GSH depletion than that evident with SNAP. In conclusion, the disruption of cellular redox homeostasis by GSH depletion leads hepatocytes to be more susceptible to NO (especially S-nitrosothiols) and subsequent necrotic cell death.

Animals↗
PubMed2004

A mitochondrial role for catabolism of nitric oxide in cardiomyocytes not involving oxymyoglobin.

The maximal concentration of nitric oxide (NO) developing in cultured cells following stimulation of endogenous NO synthases was shown to be submicromolar by NO-selective microelectrode measurements. In electron paramagnetic resonance experiments with isolated and finely divided pericardium, NO was found to react with oxymyoglobin to form metmyoglobin provided that NO was supplied at concentrations in excess of a few micromolar. However, at NO concentrations achievable by endogenous sources, this reaction did not take place to any measurable extent. Oxidative conversion of NO to nitrite ion by cytochrome c oxidase appears to be the most plausible route for cellular catabolism of NO.

Animals↗
PubMed2003

Reversal of cyanide inhibition of cytochrome c oxidase by the auxiliary substrate nitric oxide: an endogenous antidote to cyanide poisoning?

Nitric oxide (NO) is shown to overcome the cyanide inhibition of cytochrome c oxidase in the presence of excess ferrocytochrome c and oxygen. Addition of NO to the partially reduced cyanide-inhibited form of the bovine enzyme is shown by electron paramagnetic resonance spectroscopy to result in substitution of cyanide at ferriheme a3 by NO with reduction of the heme. The resulting nitrosylferroheme a3 is a 5-coordinate structure, the proximal bond to histidine having been broken. NO does not simply act as a reversibly bound competitive inhibitor but is an auxiliary substrate consumed in a catalytic cycle along with ferrocytochrome c and oxygen. The implications of this observation with regard to estimates of steady-state NO levels in vivo is discussed. Given the multiple sources of NO available to mitochondria, the present results appear to explain in part some of the curious biomedical observations reported by other laboratories; for example, the kidneys of cyanide poisoning victims surprisingly exhibit no significant irreversible damage, and lethal doses of potassium cyanide are able to inhibit cytochrome c oxidase activity by only approximately 50% in brain mitochondria.

Animals↗
PubMed2003

The case of the missing NO-hemoglobin: spectral changes suggestive of heme redox reactions reflect changes in NO-heme geometry.

When low levels of gaseous nitric oxide (NO) are equilibrated with deoxygenated Hb, all NO added can be accounted for in terms of hexacoordinate and pentacoordinate forms of NO-Hb, despite recent reports on NO disappearance from heme groups to form nitroxyl anions or S-nitrosated Hb at low ratios of NO to Hb. We demonstrate that a fraction of the spectral signature of fully nitrosylated (largely hexacoordinate) Hb disappears as the pentacoordinate state forms and reappears when pentacoordinate NO-Hb is reconverted to the hexacoordinate condition. We show that the spectral changes associated with these reversible shifts in NO- heme geometry can be remarkably well approximated as variations in the contributions from fully nitrosylated Hb and oxidized Hb (MetHb). As a result, increases in the level of pentacoordinate NO-Hb that occur at low NO to Hb ratios can be misinterpreted as increases in MetHb levels associated with NO-dependent heme oxidation. Conversely, any decrease in levels of pentacoordinate NO-Hb can be misinterpreted as a disappearance of MetHb associated with NO-dependent heme reduction. Transitions between pentacoordinate and hexacoordinate forms of NO-Hb with spectral changes suggestive of changes in levels of heme-bound NO are sensitive to the protein's quaternary conformation and can be brought about by alterations in anion levels or the degree of heme saturation with either O2 or NO.

Electron Spin Resonance Spectroscopy↗
PubMed2002

Visible region MCD and MLD spectra of nitrosylferrohemoglobin and oxyhemoglobin.

Magnetic circular dichroism (MCD) and magnetic linear dichroism (MLD) spectroscopies at various applied magnetic fields (0-6T) and temperatures (2.0-31K) have been used to investigate the electronic properties of the visible (Q(0-0), or alpha band) region of oxy- and nitrosylferrohemoglobin (HbNO). OxyHb, a d(6) (S=0) diamagnet, exhibits the expected pseudo-first derivative MCD and pseudo-second derivative MLD temperature-independent features centered at 574nm. HbNO, a d(7) (S=1/2) paramagnet, also exhibits a temperature-independent pseudo-first derivative MCD spectrum, but centered at 571nm. So far as we are aware, this behavior is unprecedented in the MCD spectra of paramagetic iron-porphyrins, which are expected to be dominated by temperature-dependent C(0) terms. The HbNO MCD spectrum does, however, demonstrate limited field-dependent saturation magnetization behavior and the MLD spectrum is currently below the detection limit. In addition, an MCD signal from reoxygenated venous blood is reported and compared with MCD signals from oxy- and HbNO derivatives. Finally, a combination of MCD and MLD spectroscopies has been used to estimate the orbital angular momentum (M(L)) value of the alpha band excited state of oxyHb as 4.2 (+/-0.7).

Circular Dichroism↗
PubMed2002

The catabolic fate of nitric oxide: the nitric oxide oxidase and peroxynitrite reductase activities of cytochrome oxidase.

Stimulation of cardiomyocytes to endogenously evolve nitric oxide is shown by microsensor measurements on single cells to lead to transient nitric oxide concentrations of a few hundred nanomolar. At these submicromolar concentrations, no evidence could be found for the expected reaction between nitric oxide generated and the oxymyoglobin present in the cells: nitric oxide + oxymyoglobin --> nitrate + metmyoglobin. No metmyoglobin formation was detected by electron paramagnetic resonance spectroscopy, and microsensor measurements revealed near quantitative conversion of the nitric oxide to nitrite rather than nitrate ion. Moreover, the rate of nitrite formation is shown to be too rapid to be accounted for by non-enzymatic means. The essentially quantitative and rapid catabolism of nitric oxide to nitrite ion can plausibly be explained on the basis of a cycle of reactions catalyzed by cytochrome c oxidase. It is demonstrated with the purified hemoproteins in vitro that the terminal oxidase can outcompete oxymyoglobin for available nitric oxide. It is proposed that under normal physiological and most pathological (non-inflammatory) conditions, reaction with cytochrome c oxidase is the major route by which NO is removed from mitochondria-rich cells.

Animals↗
PubMed2002

Multi-center retrospective evaluation of carbonyl iron ingestions.

There is limited published data concerning carbonyl iron ingestion and no data on overdose. A retrospective chart review of all patients with carbonyl iron ingestion reported to 5 regional poison information centers from January 1998 to April 2000 was done. Thirty-three patients with carbonyl iron ingestion were reported. Twenty-seven patients (82%) were managed without referral to a health care facility. The mean and median age of these patients was 3years and 20 months, respectively. The mean dose ingested was 11.2 mg/kg (range of 2.2 to 34.5 mg/kg) with no effects noted. Six patients evaluated in the emergency ingested a mean of 34 mg/kg (range of 12 to 72 mg/kg). The mean peak serum iron concentrations in 4/6 were 82 microg/dL (range of 36 to 177 microg/dL). One child with a history of flu-like symptoms reported diarrhea, fever and lethargy and had a serum iron concentration of 36 microg/dL; symptoms were believed unrelated to the exposure. In this limited first case series of iron carbonyl ingestion, serious toxicity did not occur.

Adolescent↗
PubMed1998

The Alkaline Transition of Bis(N-acetylated) Heme Undecapeptide.

Alkaline forms of the ferric bis(N-acetylated) heme undecapeptide of cytochrome c (N-ac-HUP) and some of its derivatives have been studied by electronic absorption and electron paramagnetic resonance spectroscopies. Surprisingly, even at pH >12, no evidence could be found for the formation of a hydroxyl ion adduct, in direct contrast to a previous report concerning ferric heme peptides encapsulated in detergent micelles (Mazumdar et al. Inorg. Chem. 1991, 30, 700-705). A spectroscopically determined pK(a) of approximately 9 is assigned to the deprotonation of the constituent histidine ligand of heme iron in N-ac-HUP. The present findings are not entirely in keeping with those of an earlier study concerning the properties of N-acetylated heme octapeptide (Wang et al. J. Biol. Chem. 1992, 35, 15310-15318), the differences observed being attributed to the buffering media employed in the two investigations. The implications of the current results in relation to a better understanding of the alkaline transitions observed in hemoglobins and myoglobins is considered.

Journal Article↗
PubMed1996

Characterization of N-Acetylated Heme Undecapeptide and Some of Its Derivatives in Aqueous Media: Monomeric Model Systems for Hemoproteins.

The heme undecapeptide of cytochrome c has been converted to a bis(N-acetylated) derivative by reaction with acetic anhydride. The structure of the product has been confirmed by liquid secondary-ion mass spectrometry. As anticipated, the N-acetylated molecule exhibits much less tendency to aggregate in aqueous solution than its heme undecapeptide precursor. Around neutral pH, one axial ligand on the heme iron is provided by the same histidine residue as in the native cytochrome. The other axial ligand can be varied by the addition of exogenous donor species to produce a range of hemoprotein model compounds exhibiting mixed axial ligation. Contrary to the findings of Othman et al. [Biochemistry 1994, 33, 15437-15448] concerning heme octapeptide, the N-acetylated undecapeptide showed no tendency to bind more than one exogenous ligand per heme. At concentrations approaching millimolar and in the absence of exogenous ligands, the N-acetylated molecule may either be monodispersed, exhibiting a characteristic high-spin (S = (5)/(2)) ferric heme electron paramagnetic resonance (EPR) signal, or exist in an EPR-silent and presumably aggregated form. Interestingly, the system displays a novel dependence on the buffer with regard to which of these two forms is present in a given sample. There is no evidence in any of the spectra for the existence of an intermediate-spin (S = (3)/(2)) ferric heme as suggested by Wang and Van Wart [J. Phys. Chem. 1989, 93, 7925-7931] to be present in aqueous solutions of N-acetylated heme octapeptide. Also, in contrast to another earlier report concerning the underivatized undecapeptide [Clore et al. Inorg. Chim. Acta 1981, 56, 143-148], the N-acetylated molecule showed no evidence of catalase activity. In fact, the heme chromophore was surprisingly unstable in the presence of hydrogen peroxide.

Journal Article↗