scholarly journals Activation of Cytochrome C Peroxidase Function Through Coordinated Foldon Loop Dynamics upon Interaction with Anionic Lipids

2021 ◽  
Author(s):  
Mingyue Li ◽  
Wanyang Sun ◽  
Vladimir A. Tyurin ◽  
Maria DeLucia ◽  
Jinwoo Ahn ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Peroxidase Activity ◽  
Magic Angle ◽  
List Type ◽  
Mitochondrial Apoptosis ◽  
Chemical Denaturation ◽  
Cyt C ◽  
Anionic Lipids ◽  
Lipid Peroxidase ◽  
Peroxidase Activation

ABSTRACTCardiolipin (CL) is a mitochondrial anionic lipid that plays important roles in the regulation and signaling of mitochondrial apoptosis. CL peroxidation catalyzed by the assembly of CL-cytochrome c (cyt c) complexes at the inner mitochondrial membrane is a critical checkpoint. The structural changes in the protein, associated with peroxidase activation by CL and different anionic lipids, are not known at a molecular level. To better understand these peripheral protein-lipid interactions, we compare how phosphatidylglycerol (PG) and CL lipids trigger cyt c peroxidase activation, and correlate functional differences to structural and motional changes in membrane-associated cyt c. Structural and motional studies of the bound protein are enabled by magic angle spinning solid state NMR spectroscopy, while lipid peroxidase activity is assayed by mass spectrometry. PG binding results in a surface-bound state that preserves a nativelike fold, which nonetheless allows for significant peroxidase activity, though at a lower level than binding its native substrate CL. Lipid-specific differences in peroxidase activation are found to correlate to corresponding differences in lipid-induced protein mobility, affecting specific protein segments. The dynamics of omega loops C and D are upregulated by CL binding, in a way that is remarkably controlled by the protein:lipid stoichiometry. In contrast to complete chemical denaturation, membrane-induced protein destabilization reflects a destabilization of select cyt c foldons, while the energetically most stable helices are preserved. Our studies illuminate the interplay of protein and lipid dynamics in the creation of lipid peroxidase-active proteolipid complexes implicated in early stages of mitochondrial apoptosis.GRAPHICAL ABSTRACTHIGHLIGHTSA mitochondrial protein-lipid complex regulates lipid peroxidation in apoptosis.Peroxidase-active lipid-cytochrome c complexes are reconstituted in vitro.Phosphatidylglycerol lipids are less effective activators than cardiolipin.Activity correlates to localized dynamics, distinct from chemical denaturation.A dynamic interplay of cytochrome c foldons and anionic lipids regulate activity.

scholarly journals Lysine carbonylation is a previously unrecognized contributor to peroxidase activation of cytochrome c by chloramine-T

2019 ◽  
Vol 10 (8) ◽  
pp. 2349-2359 ◽  
Author(s):  
Victor Yin ◽  
Safee H. Mian ◽  
Lars Konermann
Keyword(s):  
Cytochrome C ◽  
Ion Mobility ◽  
Peroxidase Activity ◽  
Tandem Ms ◽  
Chloramine T ◽  
Peroxidase Activation

Ion mobility-assisted tandem MS uncovers hitherto overlooked modifications that are critical for the peroxidase activity of chloramine T-modified cytochrome c.

scholarly journals Characterization of oxidation of glutathione by cytochrome c

2021 ◽  
Author(s):  
K. B. Csomó ◽  
B. Alasztics B ◽  
A. P. Sándor ◽  
A. A. Belik ◽  
G. Varga ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Respiratory Chain ◽  
Oxygen Electrode ◽  
Oxidize Glutathione ◽  
Mitochondrial Oxygen Consumption ◽  
Anionic Phospholipids ◽  
Cyt C ◽  
Membrane Bound ◽  
Anionic Lipids

AbstractCytochrome c is a member of the respiratory chain of the mitochondria. Non-membrane-bound (free) cytochrome c can be reduced by gluthatione as well as ascorbic acid. We investigated the effect of pH, Ca2+, Mg2+ and anionic phospholipids on the reduction of cytochrome c by glutathione.The reduction of cytochrome c by thiols was measured using photometry. Mitochondrial oxygen consumption was detected by use of oxygen electrode. Glutathione does not reduce cytochrome c at pH = 7.0 in the absence of Ca2+ and Mg2+. The reduction of cytochrome c by glutathione is inhibited by anionic lipids, especially cardiolipin. The typical conditions of apoptosis—elevated pH, Ca2+ level and Mg2+—increases the reduction of cytochrome c. Glutathione (5 mM) causes increased mitochondrial O2 consumption at pH = 8.0, in the presence of ADP either 1 mM Mg2+ or 1 mM Ca2+. Our results suggest that membrane bound cyt c does not oxidize glutathione. Free (not membrane bound) cytochrome c can oxidize glutathione. In mitochondria, O2 is depleted only in the presence of ADP, so the O2 depletion observed in the presence of glutathione can be related to the respiratory chain. Decreased glutathione levels play a role in apoptosis. Therefore, membrane unbound cyt c can contribute to apoptosis by oxidation of glutathione.

Effects of cytochrome c on the mitochondrial apoptosis-induced channel MAC

2004 ◽  
Vol 286 (5) ◽  
pp. C1109-C1117 ◽  
Author(s):  
Liang Guo ◽  
Dawn Pietkiewicz ◽  
Evgeny V. Pavlov ◽  
Sergey M. Grigoriev ◽  
John J. Kasianowicz ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Outer Membrane ◽  
Cell Types ◽  
Rnase A ◽  
Mitochondrial Outer Membrane ◽  
Dependent Manner ◽  
Mitochondrial Apoptosis ◽  
Noise Type ◽  
Voltage Dependent

Recent studies indicate that cytochrome c is released early in apoptosis without loss of integrity of the mitochondrial outer membrane in some cell types. The high-conductance mitochondrial apoptosis-induced channel (MAC) forms in the outer membrane early in apoptosis of FL5.12 cells. Physiological (micromolar) levels of cytochrome c alter MAC activity, and these effects are referred to as types 1 and 2. Type 1 effects are consistent with a partitioning of cytochrome c into the pore of MAC and include a modest decrease in conductance that is dose and voltage dependent, reversible, and has an increase in noise. Type 2 effects may correspond to “plugging” of the pore or destabilization of the open state. Type 2 effects are a dose-dependent, voltage-independent, and irreversible decrease in conductance. MAC is a heterogeneous channel with variable conductance. Cytochrome c affects MAC in a pore size-dependent manner, with maximal effects of cytochrome c on MAC with conductance of 1.9–5.4 nS. The effects of cytochrome c, RNase A, and high salt on MAC indicate that size, rather than charge, is crucial. The effects of dextran molecules of various sizes indicate that the pore diameter of MAC is slightly larger than that of 17-kDa dextran, which should be sufficient to allow the passage of 12-kDa cytochrome c. These findings are consistent with the notion that MAC is the pore through which cytochrome c is released from mitochondria during apoptosis.

Expression of Cyt-c-Mediated Mitochondrial Apoptosis-Related Proteins in Rat Renal Proximal Tubules during Development

Nephron ◽  
2016 ◽  
Vol 135 (1) ◽  
pp. 77-86 ◽  
Author(s):  
Xiao-Feng Song ◽  
He Tian ◽  
Ping Zhang ◽  
Zhen-Xing Zhang
Keyword(s):  
Proximal Tubules ◽  
Mitochondrial Apoptosis ◽  
Renal Proximal Tubules ◽  
Cyt C ◽  
Related Proteins

AB0135 THE MITOCHONDRIAL-RETICULAR NETWORK (MRN) OF NEUTROPHILIC LEUKOCYTES OF SYNOVIAL FLUID (SF) OF PATIENTS WITH SLE AND RA

2020 ◽  
Vol 79 (Suppl 1) ◽  
pp. 1368.1-1368
Author(s):  
G. V. Kudriavtseva ◽  
Y. Malenkov ◽  
V. Shishkin ◽  
V. V. Shishkin
Keyword(s):  
Free Radicals ◽  
Cytochrome C ◽  
Electrophoretic Mobility ◽  
Oxygen Free Radicals ◽  
Electrokinetic Potential ◽  
Active Forms Of Oxygen ◽  
Autoimmune Pathology ◽  
Neutrophilic Leukocytes ◽  
Swelling Rate ◽  
Cyt C

Background:Objectives:It has been established that in cells, in particular in neutrophilic leukocytes of SF, mitochondria form a mitochondrial-reticular dynamic spatial network (MRN). MRN is the epicenter of apoptosis, reflecting structural and functional changes in the immuno-complex pathology in SLE and RA.Methods:SF was analyzed in patients: 10 SLE (43 ± 2.3 years), 13 RA (45 ± 1.6 years) and 8 donors (42 ± 3.7 years, postmortem). Neutrophilic leukocytes from the SF were isolated by standard methods and resuspended in a composition medium: 70 mM NaCl; 140 mM sucrose; 5.6 mM KCl; 10 mM pyruvate; 8 mM MOPS; pH = 7.4. The cell suspension was centrifuged for 5 min at 800 g. MRN was isolated by centrifuging the resulting supernatant for 15 min at 12 000 g. The resulting MRN fragments were resuspended in citrate-phosphate buffer (pH = 7.4) and used in experiments. The activity of adenosine monophosphate-activated protein kinase (AMPK) was evaluated by Western blotting. Quantitative determination of cytochrome C (Cyt C) was carried out by enzyme immunoassay method using the Human Cytochrome c Platinum ELISA kit (eBioscience, USA). Active forms of oxygen free radicals (AFRF) were registered by EPR. The swelling rate of MRN fragments was determined spectrophotometrically at 540 nm. The electrophoretic mobility (EM) of MRN fragments was determined by the automatic microscope “Parmoquant-2”.Results:MRN of neutrophilic leukocytes of the SF undergoes significant adaptive rearrangements during the development of SLE and RA (tab.1). On average, the expression of biochemical indicators of autophagy (AMPK), apoptosis (Сyt. C), necrosis (level of oxygen free radicals, low-amplitude swelling rate) increases by 2-3 times compared with the conventional norm. Particular attention should be paid to pathological changes in the electrokinetic potential of MRN, which determines the functional state of the SF as a whole as a colloidal system. Obviously, in SLE and RA, depletion of the energy of MRN (a sharp increase in the activity of AMRK), activation of free radical processes, disruption of intracellular ion homeostasis due to an increase in the rate of swelling of MRN as a manifestation of a compensatory-adaptive reaction. It ultimately leads to a decrease in electrokinetic properties of MRN. Thus EM is an integral indicator of physico-chemical properties and architectonics of MRN pointihg to the development of autoimmune pathology.Table 1.EXPRESSION OF INDUCTORS OF AUTOPHAGY, APOPTOSIS, NECROSIS AND ELECTROPHORETIC MOBILITY OF MRN FRAGMENTS OF NEUTROPHILIC LEUKOCYTES OF SF IN SLE AND RAExperience TermsAMPK,cond.unit/mg proteinCyt C,ng/mlAFRF, unit/mg proteinSwelling rate of MRN,/min ·mg proteinEM,m/v · secDonor (8)0,51±0,0523,7±5,47,3±2,40,177± 0,0041,58± 0,07SLE (10)1,73±0,04**49,3±6,5*21,3±5,1**0,435±0,005***0,35±0,05***RA (13)1,25±0,07**47,8±4.8*15,7±4,3*0,410±0,007***0,41±0,07***Notes: differences with the control norm: * - p <0.05; ** - p <0.01; *** - p <0.001.Conclusion:Endoplasmic stress occurs in SF cells during the development of SLE and RA, blocking of autophagy and apoptosis leads to a breakdown of neutrophilic leukocyte MRN, accumulation of high molecular products of tissue decay - phlogogens in the intercellular space, among which the expression in the context is characterized by proteins - chaperones Hsp 60-100. These processes are accompanied by a shift in the bioelectric homeostasis of MRN neutrophilic leukocytes, an increase in their swelling rate and a significant decrease in their electrokinetic potential. The described MRN reactions of neutrophilic leukocytes of the SF should be taken into account when developing pharmacologically induced apoptosis as a new approach in the treatment of autoimmune diseasesReferences:[1]Shishkin V. I. et al. Ann Rheum Dis 2017; 76: No 6, p.1077-1078; DOI:10.1136/annrheumdis-2017-eular5364Disclosure of Interests:None declared

scholarly journals Structure of a mitochondrial cytochrome c conformer competent for peroxidase activity

2014 ◽  
Vol 111 (18) ◽  
pp. 6648-6653 ◽  
Author(s):  
L. J. McClelland ◽  
T.-C. Mou ◽  
M. E. Jeakins-Cooley ◽  
S. R. Sprang ◽  
B. E. Bowler
Keyword(s):  
Cytochrome C ◽  
Peroxidase Activity ◽  
Mitochondrial Cytochrome ◽  
Mitochondrial Cytochrome C

The proportion of Met80-sulfoxide dictates peroxidase activity of human cytochrome c

2018 ◽  
Vol 47 (27) ◽  
pp. 9128-9135 ◽  
Author(s):  
Rinky D. Parakra ◽  
Torsten Kleffmann ◽  
Guy N. L. Jameson ◽  
Elizabeth C. Ledgerwood
Keyword(s):  
Cytochrome C ◽  
Peroxidase Activity ◽  
Human Cytochrome ◽  
Human Cytochrome C

Peroxidase activity of cytochrome c is activated and deactivated by methionine 80 oxidation to the sulfoxide and sulfone respectively.

Differences in renal metabolism of insulin and cytochrome c

1988 ◽  
Vol 254 (4) ◽  
pp. E419-E428 ◽  
Author(s):  
J. Herrman ◽  
R. E. Simmons ◽  
B. H. Frank ◽  
R. Rabkin
Keyword(s):  
Cytochrome C ◽  
Lysosomal Enzyme ◽  
Subcellular Distribution ◽  
Sucrose Gradient ◽  
Insulin Degradation ◽  
Renal Metabolism ◽  
Small Proteins ◽  
Cyt C ◽  
Alternate Routes

Kidneys degrade small proteins such as cytochrome c (CYT c) by the classic lysosomal pathway. However, because alternate routes for the transport and degradation of protein hormones have been identified in other tissues, we set out to determine whether extralysosomal sites might participate in the renal degradation of insulin. First, we compared the effect of the lysosomal inhibitor NH4Cl on insulin and CYT c degradation by isolated perfused rat kidneys. After kidneys were loaded with radiolabeled proteins to allow for absorption and transport to lysosomes, degradation was measured in the presence or absence of inhibitors. Control kidneys degraded 45 +/- 1.5% of the trapped CYT c per hour, and this was inhibited 62 +/- 1.3% by NH4Cl. In contrast, 86 +/- 2.4% of the trapped insulin was degraded per hour, and this was inhibited 26 +/- 4% by NH4Cl. Next we followed the subcellular distribution of 125I-labeled insulin in kidneys exposed to 125I-labeled insulin in vivo or when isolated and perfused. Under both circumstances the distribution of insulin on a linear sucrose gradient differed from that of the lysosomal enzyme N-acetyl-beta-glucosaminidase. In contrast, [14CH3]CYT c, injected in vivo, distributed over a density similar to the lysosomal marker. Thus important differences exist between the renal metabolism of CYT c, which proceeds in lysosomes, and the renal metabolism of insulin. These include rate of degradation, sensitivity to NH4Cl, and subcellular sites of localization. Accordingly, we suggest that insulin degradation may occur, at least in part, in a different compartment from the classic lysosomal site of protein degradation.

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