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1.
We studied the activity of a calcium-dependent proteinase, calpain, in subcellular fractions obtained from rat brain tissue. The rates of calpain-mediated hydrolysis of fluorescein isothiocyanate (FITC)-labeled substrates, casein and fodrin, were comparable; in the former case the rate was higher. This fact stipulated the choice of fluorescent-labeled casein as an adequate substrate. The greatest enzyme activity of calpain (87% of total) was found in the cytoplasmic fraction. At the same time, quite detectable enzyme activities were observed in the investigated membrane fractions obtained from rat brain tissue (coarse mitochondrial fraction, microsomes, and myelin). The highest specific calpain activity was registered in the cytoplasmic fraction. The enzyme activity was efficiently suppressed in the presence of calpain inhibitor I and increased after purification of the preparations from an endogenous calpain inhibitor, calpastatin.Neirofiziologiya/Neurophysiology, Vol. 36, No. 4, pp. 265–271, July–August, 2004.This revised version was published online in April 2005 with a corrected cover date. 相似文献
2.
Ramunas Rolius Chloe Antoniou Lidia A. Nazarova Stephen H. Kim Garrett Cobb Pooja Gala Priyanka Rajaram Qufei Li Leslie W.-M. Fung 《Cellular & molecular biology letters》2010,15(3):395-405
Calpains and caspases are ubiquitous cysteine proteases that are associated with a variety of cellular pathways. Calpains
are involved in processes such as long term potentiation, cell motility and apoptosis, and have been shown to cleave non-erythroid
(brain) α- and β-spectrin and erythroid β-spectrin. The cleavage of erythroid α-spectrin by calpain has not been reported.
Caspases play an important role in the initiation and execution of apoptosis, and have been shown to cleave non-erythroid
but not erythroid spectrin. We have studied the effect of spectrin fragments on calpain and caspase activities. The erythroid
and non-erythroid spectrin fragments used were from the N-terminal region of α-spectrin, and C-terminal region of β-spectrin,
both consisting of regions involved in spectrin tetramer formation. We observed that the all spectrin fragments exhibited
a concentration-dependent inhibitory effect on calpain, but not caspase activity. It is clear that additional studies are
warranted to determine the physiological significance of calpain inhibition by spectrin fragments. Our findings suggest that
calpain activity is modulated by the presence of spectrin partial domains at the tetramerization site. It is not clear whether
the inhibitory effect is substrate specific or is a general effect. Further studies of this inhibitory effect may lead to
the identification and development of new therapeutic agents specifically for calpains, but not for caspases. Proteins/peptides
with a coiled coil helical conformation should be studied for potential inhibitory effects on calpain activity. 相似文献
3.
Giguere CJ Covington MD Schnellmann RG 《Biochemical and biophysical research communications》2008,366(1):258-262
Calpains, Ca2+-activated cysteine proteases, have been implicated in the progression of multiple disease states. We recently identified calpain 10 as a mitochondrial calpain that is involved in Ca2+-induced mitochondrial dysfunction. The goals of this study were to characterize the expression and activity of renal mitochondrial calpain 10 in rabbit, mouse, and rat. Using shRNA technology and immunoblot analysis three previously postulated splice variants of calpain 10 were identified (50, 56, and 75 kDa). SLLVY-AMC zymography and immunoblot analysis was used to directly link calpeptin-sensitive calpain activity to calpain 10 splice variants. Rabbit, mouse, and rat kidney mitochondria contained 75 kDa (calpain 10a), 56 kDa (calpain 10c or 10d), and 50 kDa (calpain 10e) splice variants. Interestingly, zymography yielded distinct bands of calpain activity containing multiple calpain 10 splice variants in all species. These results provide evidence that several previously postulated splice variants of calpain 10 are localized to the mitochondria in kidneys of rabbits, rats, and mice. 相似文献
4.
Koszela-Piotrowska I Choma K Bednarczyk P Dołowy K Szewczyk A Kunz WS Malekova L Kominkova V Ondrias K 《Cellular & molecular biology letters》2007,12(4):493-508
Ion channels selective for chloride ions are present in all biological membranes, where they regulate the cell volume or membrane
potential. Various chloride channels from mitochondrial membranes have been described in recent years. The aim of our study
was to characterize the effect of stilbene derivatives on single-chloride channel activity in the inner mitochondrial membrane.
The measurements were performed after the reconstitution into a planar lipid bilayer of the inner mitochondrial membranes
from rat skeletal muscle (SMM), rat brain (BM) and heart (HM) mitochondria. After incorporation in a symmetric 450/450 mM
KCl solution (cis/trans), the chloride channels were recorded with a mean conductance of 155 ± 5 pS (rat skeletal muscle) and 120 ± 16 pS (rat brain).
The conductances of the chloride channels from the rat heart mitochondria in 250/50 mM KCl (cis/trans) gradient solutions were within the 70–130 pS range. The chloride channels were inhibited by these two stilbene derivatives:
4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) and 4-acetamido-4′-isothiocyanostilbene-2,2′-disulfonic acid (SITS).
The skeletal muscle mitochondrial chloride channel was blocked after the addition of 1 mM DIDS or SITS, whereas the brain
mitochondrial channel was blocked by 300 μM DIDS or SITS. The chloride channel from the rat heart mitochondria was inhibited
by 50–100 μM DIDS. The inhibitory effect of DIDS was irreversible. Our results confirm the presence of chloride channels sensitive
to stilbene derivatives in the inner mitochondrial membrane from rat skeletal muscle, brain and heart cells. 相似文献
5.
Neuronal Mitochondrial Toxicity of Malondialdehyde: Inhibitory Effects on Respiratory Function and Enzyme Activities in Rat Brain Mitochondria 总被引:1,自引:0,他引:1
Jiangang Long Changsheng Liu Lijuan Sun Hongxiang Gao Jiankang Liu 《Neurochemical research》2009,34(4):786-794
Malondialdehyde (MDA) is a product of oxidative damage to lipids, amino acids and DNA, and accumulates with aging and diseases.
MDA can possibly react with amines so as to modify proteins and inactivate enzymes; it can also modify nucleosides so as to
cause mutagenicity. Brain mitochondrial dysfunction is a major contributor to aging and neurodegenerative diseases. We hypothesize
that MDA accumulated during aging targets mitochondrial enzymes so as to cause further mitochondrial dysfunction and additional
contributions to aging and neurodegeneration. Herein, we investigated the neuronal mitochondrial toxic effects of MDA on mitochondrial
respiration and activities of enzymes (mitochondrial complexes I–V, α-ketoglutarate dehydrogenase (KGDH) and pyruvate dehydrogenase
(PDH)), in isolated rat brain mitochondria. MDA depressed mitochondrial membrane potential, and also showed a dose-dependent
inhibition of mitochondrial complex I- and complex II-linked respiration. Complex I and II, and PDH activities were depressed
by MDA at ≥0.2 μmol/mg; KGDH and complex V were inhibited by ≥0.4 and ≥1.6 μmol MDA/mg, respectively. However, MDA did not
have any toxic effects on complex III and IV activities over the range 0–2 μmol/mg. MDA significantly elevated mitochondrial
reactive oxygen species (ROS) and protein carbonyls at 0.2 and 0.002 μmol/mg, respectively. As for the antioxidant defense
system, a high dose of MDA slightly decreased mitochondrial GSH and superoxide dismutase. These results demonstrate that MDA
causes neuronal mitochondrial dysfunction by directly promoting generation of ROS and modifying mitochondrial proteins. The
results suggest that MDA-induced neuronal mitochondrial toxicity may be an important contributing factor to brain aging and
neurodegenerative diseases.
Special issue article in honor of Dr. Akitane Mori. 相似文献
6.
It was recently asserted that the voltage-dependent anion channel (VDAC) serves as a global regulator, or governor, of mitochondrial
function (Lemasters and Holmuhamedov, Biochim Biophys Acta 1762:181–190, 2006). Indeed, VDAC, positioned on the interface between mitochondria and the cytosol (Colombini, Mol Cell Biochem 256:107–115, 2004), is at the control point of mitochondria life and death. This large channel plays the role of a “switch” that defines in
which direction mitochondria will go: to normal respiration or to suppression of mitochondria metabolism that leads to apoptosis
and cell death. As the most abundant protein in the mitochondrial outer membrane (MOM), VDAC is known to be responsible for
ATP/ADP exchange and for the fluxes of other metabolites across MOM. It controls them by switching between the open and “closed”
states that are virtually impermeable to ATP and ADP. This control has dual importance: in maintaining normal mitochondria
respiration and in triggering apoptosis when cytochrome c and other apoptogenic factors are released from the intermembrane
space into the cytosol. Emerging evidence indicates that VDAC closure promotes apoptotic signals without direct involvement
of VDAC in the permeability transition pore or hypothetical Bax-containing cytochrome c permeable pores. VDAC gating has been
studied extensively for the last 30 years on reconstituted VDAC channels. In this review we focus exclusively on physiologically
relevant regulators of VDAC gating such as endogenous cytosolic proteins and mitochondrial lipids. Closure of VDAC induced
by such dissimilar cytosolic proteins as pro-apoptotic tBid and dimeric tubulin is compared to show that the involved mechanisms
are rather distinct. While tBid mostly modulates VDAC voltage gating, tubulin blocks the channel with the efficiency of blockage
controlled by voltage. We also discuss how characteristic mitochondrial lipids, phospatidylethanolamine and cardiolipin, could
regulate VDAC gating. Overall, we demonstrate that VDAC gating is not just an observation made under artificial conditions
of channel reconstitution but is a major mechanism of MOM permeability control. 相似文献
7.
Upon apoptosis induction, the proapoptotic protein Bax is translocated from the cytosol to mitochondria, where it promotes release of cytochrome c, a caspase‐activating protein. However, the molecular mechanisms by which Bax triggers cytochrome c release are unknown. Here we report that before the initiation of apoptotic execution by etoposide or staurosporin, an active calpain activity cleaves Bax at its N‐terminus, generating a potent proapoptotic 18‐kDa fragment (Bax/p18). Both the calpain‐mediated Bax cleavage activity and the Bax/p18 fragment were found in the mitochondrial membrane‐enriched fraction. Cleavage of Bax was followed by release of mitochondrial cytochrome c, activation of caspase‐3, cleavage of poly(ADP‐ribose) polymerase, and fragmentation of DNA. Unlike the full‐length Bax, Bax/p18 did not interact with the antiapoptotic Bcl‐2 protein in the mitochondrial fraction of drug‐treated cells. Pretreatment with a specific calpain inhibitor calpeptin inhibited etoposide‐induced calpain activation, Bax cleavage, cytochrome c release, and caspase‐3 activation. In contrast, transfection of a cloned Bax/p18 cDNA into multiple human cancer cell lines targeted Bax/p18 to mitochondria, which was accompanied by release of cytochrome c and induction of caspase‐3‐mediated apoptosis that was not blocked by overexpression of Bcl‐2 protein. Therefore, Bax/p18 has a cytochrome c–releasing activity that promotes cell death independent of Bcl‐2. Finally, Bcl‐2 overexpression inhibited etoposide‐induced calpain activation, Bax cleavage, cytochrome c release, and apoptosis. Our results suggest that the mitochondrial calpain plays an essential role in apoptotic commitment by cleaving Bax and generating the Bax/p18 fragment, which in turn mediates cytochrome c release and initiates the apoptotic execution. J. Cell. Biochem. 80:53–72, 2000. © 2000 Wiley‐Liss, Inc. 相似文献
8.
Upon apoptosis induction, the proapoptotic protein Bax is translocated from the cytosol to mitochondria, where it promotes release of cytochrome c, a caspase-activating protein. However, the molecular mechanisms by which Bax triggers cytochrome c release are unknown. Here we report that before the initiation of apoptotic execution by etoposide or staurosporin, an active calpain activity cleaves Bax at its N-terminus, generating a potent proapoptotic 18-kDa fragment (Bax/p18). Both the calpain-mediated Bax cleavage activity and the Bax/p18 fragment were found in the mitochondrial membrane-enriched fraction. Cleavage of Bax was followed by release of mitochondrial cytochrome c, activation of caspase-3, cleavage of poly(ADP-ribose) polymerase, and fragmentation of DNA. Unlike the full-length Bax, Bax/p18 did not interact with the antiapoptotic Bcl-2 protein in the mitochondrial fraction of drug-treated cells. Pretreatment with a specific calpain inhibitor calpeptin inhibited etoposide-induced calpain activation, Bax cleavage, cytochrome c release, and caspase-3 activation. In contrast, transfection of a cloned Bax/p18 cDNA into multiple human cancer cell lines targeted Bax/p18 to mitochondria, which was accompanied by release of cytochrome c and induction of caspase-3-mediated apoptosis that was not blocked by overexpression of Bcl-2 protein. Therefore, Bax/p18 has a cytochrome c-releasing activity that promotes cell death independent of Bcl-2. Finally, Bcl-2 overexpression inhibited etoposide-induced calpain activation, Bax cleavage, cytochrome c release, and apoptosis. Our results suggest that the mitochondrial calpain plays an essential role in apoptotic commitment by cleaving Bax and generating the Bax/p18 fragment, which in turn mediates cytochrome c release and initiates the apoptotic execution. 相似文献
9.
Functional processes in freshwater ecosystems are highly influenced by acidic conditions. Foodwebs are affected and macroinvertebrate
species diversity is decreased. This study aims to investigate leaf decomposition at very low pH in the acidic Banyupahit–Banyuputih
river originating from the acidic crater lake Kawah Ijen in Indonesia. Leaf decomposition experiments were carried out for
200 days in the acidic river at pHs of approximately 0.7, 2.3 and 3.0 and in the neutral Kali Sengon river, using leaves from
teak, Tectona grandis, and bamboo, Bambusa sp. Two different types of leaf packs were used: fine mesh size packs were used to exclude macroinvertebrates and coarse
mesh size packs allowed macroinvertebrate colonization. Clear differences in decomposition rate were observed between the
neutral Kali Sengon and the acidic Banyupahit–Banyuputih river with decomposition in the Kali Sengon river proceeding significantly
faster for both leaf types. In the Kali Sengon k values (d−1) over 46 days were 0.0202 for fine teak, 0.0236 for coarse teak, 0.0114 for fine bamboo and 0.0151 for coarse bamboo. No
significant differences were observed between the three sites in the acidic Banyupahit–Banyuputih river with k values of 0.0034–0.0066 for fine teak, 0.0002–0.0057 for coarse teak, 0.0029–0.0054 for fine bamboo and 0.0000–0.0068 for
coarse bamboo. Moreover, no clear adaptation of macroinvertebrates or microbes to low pH conditions could be detected. The
coarse mesh leaf packs in the neutral Kali Sengon river revealed that macroinvertebrates are important in the breakdown process.
Fine mesh packs revealed that microbial activity is depressed under acidic conditions. Based on this evidence, we conclude
that the toxicity at low pH conditions, and probably also the precipitation of metals on the leaf material, seriously affects
leaf decomposition. 相似文献
10.
Decomposition and carbon cycling of dead trees in tropical forests of the central Amazon 总被引:11,自引:0,他引:11
Decomposition rate constants were measured for boles of 155 large dead trees (>10 cm diameter) in central Amazon forests.
Mortality data from 21 ha of permanent inventory plots, monitored for 10–15 years, were used to select dead trees for sampling.
Measured rate constants varied by over 1.5 orders of magnitude (0.015–0.67 year–1), averaging 0.19 year–1 with predicted error of 0.026 year. Wood density and bole diameter were significantly and inversely correlated with rate
constants. A tree of average biomass was predicted to decompose at 0.17 year–1. Based on mortality data, an average of 7.0 trees ha–1 year–1 died producing 3.6 Mg ha–1 year–1 of coarse litter (>10 cm diameter). Mean coarse litter standing-stocks were predicted to be 21 Mg ha–1, with a mean residence time of 5.9 years, and a maximum mean carbon flux to the atmosphere of 1.8 Mg C ha–1 year–1. Total litter is estimated to be partitioned into 16% fine wood, 30% coarse wood, and 54% non-woody litter (e.g., leaves,
fruits, flowers). Decomposition rate constants for coarse litter were compiled from 20 globally distributed studies. Rates
were highly correlated with mean annual temperature, giving a respiration quotient (Q
10) of 2.4 (10°C–1).
Received: 14 June 1999 / Accepted: 31 August 1999 相似文献
11.
Harry Walter Eugene J. Krob Rosalinda B. Wenby Herbert J. Meiselman 《Cell biochemistry and biophysics》1990,16(3):139-148
Rats were injected with59Fe-ferrous citrate and bled thereafter at different times (16 h to 49 d). This gave rise to red cell populations in which
cells corresponding in age to the time elapsed between injection and bleeding were labeled. The anticoagulant used was either
acid-citrate-dextrose (ACD) with a pH adjusted to 7.3 or ACD (pH 5.1). Final pH of the collected blood was about 7.2–7.4 in
the former case and 6.4–6.7 in the latter. Red cells were then centrifuged (5) and approximately 7–10% of the packed cells
from the top and 7–10% from the bottom of the cell column collected. When reticulocytes are the predominant labeled red cell
population, as in blood obtained for about 24 h after isotope injection, a fractionation of these cells and mature erythrocytes
is in evidence only when blood is collected at the higher pH. Thus, at pH 7.2–7.4 ratios of specific radioactivities of cells
in top fraction/cells in an unfractionated sample are about 3, whereas at pH 6.4–6.7, the analogous ratios are 1 or less.
These differences in specific activity ratios, as a function of pH at collection, virtually disappear after about 4 d following
isotope injection.
The lower pH is known to increase the volume and decrease the density of mature red blood cells. The marked effect of pH on
cellular fractionation could be correlated with the smaller change in rat reticulocyte density and volume in acid medium.
At pH 6.4–6.7, the densities of mature erythrocytes and reticulocytes are so close that their physical separation by centrifugation
is not feasible. 相似文献
12.
Effect of Lyophilization and Freeze-thawing on the Stability of siRNA-liposome Complexes 总被引:1,自引:0,他引:1
The purpose of this research was to describe the application of lyophilization in the delivery of siRNA using cationic lipids
by addressing the long-term formulation/stability issues associated with cationic lipids and to understand the mechanism of
lyoprotection. siRNA liposomes complexes were formed in different potential cyro/lyoprotectants and subjected to either lyophilization
or freeze thaw cycles. siRNA, liposomes and/or lipoplexes were tested for activity, SYBR Green I binding, cellular uptake
and particle size. The lipoplexes when lyophilized in the presence of sugars as lyoprotectants could be lyophilized and reconstituted
without loss of transfection efficacy but in ionic solutions they lost 65–75% of their functionality. The mechanism of this
loss of activity was further investigated. The lyophilization process did not alter siRNA’s intrinsic biological activity
as was evident by the ability of lyophilized siRNA to retain functionality and SYBR green I binding ability. While the lipoplex
size dramatically increased (∼50–70 times) after lyophilization in the absence of non-ionic lyoprotectants. This increase
in size correlated to the decrease in cellular accumulation of siRNA and a decrease in activity. In conclusion, siRNAs can
be applied in cationic lipid lyophilized formulations and these complexes represent a potential method of increasing the stability
of pre-formed complex. 相似文献
13.
Kinetics of degradation of labelled proteins was followed in two asporogenic mutants ofBacillus megaterium during incubation in a sporulation medium. Both the mutant producing exocellular protease (KM 1prn
+) and the mutant not producing the enzyme (KM 12prn) were found to contain a labile protein fraction, whose proportion decreases with prolonged time of labelling and whose half-life
is about 1 h. Most proteins were relatively stable and were degraded at a rate of 1 %/h and 2 %/h in strains KM 1 and KM 12,
respectively (half life 70–80 h and 35–40 h in strains KM 1 and KM 12, respectively). The intracellular proteolytic activity
of the KM 12 mutant remains practically the same during incubation in the sporulation medium or slowly increases. The labile
protein fraction practically disappears from the cells after a 3.5-h incubation. When such a culture is then subjected to
a shift-up and transferred again to the sporulation medium, the rate of protein turnover temporarily increases. The temporary
increase of the turnover rate is caused by a partial replenishment of the labile protein fraction rather than by an accelerated
degradation of the relatively stable fraction. The intracellular proteolytic activity does not increase under these conditions.
The wild sporogenic strain ofB. megaterium also contains the labile protein fraction. Its half protein life is 1 h or less. However, the second protein fraction is
degraded much more rapidly than in the asporogenic mutants and its half life is 6–7 h. 相似文献
14.
Saehun Mun Eric A. Decker Yeonhwa Park Jochen Weiss D. Julian McClements 《Food biophysics》2006,1(1):21-29
The objective of this study was to investigate the influence of interfacial composition and electrical charge on the in vitro digestion of emulsified fats by pancreatic lipase. An electrostatic layer-by-layer deposition technique was used to prepare corn oil-in-water emulsions (3 wt% oil) that contained droplets coated by (1) lecithin, (2) lecithin–chitosan, or (3) lecithin–chitosan–pectin. Pancreatic lipase (1.6 mg mL−1) and/or bile extract (5.0 mg mL−1) were added to each emulsion, and the particle charge, droplet aggregation, and free fatty acids released were measured. In the presence of bile extract, the amount of fatty acids released per unit amount of emulsion was much lower in the emulsions containing droplets coated by lecithin–chitosan (38 ± 16 μmol mL−1) than those containing droplets coated by lecithin (250 ± 70 μmol mL−1) or lecithin–chitosan–pectin (274 ± 80 μmol mL−1). In addition, there was much more extensive droplet aggregation in the lecithin–chitosan emulsion than in the other two emulsions. We postulated that lipase activity was reduced in the lecithin–chitosan emulsion as a result of the formation of a relatively thick cationic layer around each droplet, as well as the formation of large flocs, which restricted the access of the pancreatic lipase to the lipids within the droplets. Our results also suggest that droplets initially coated by a lecithin–chitosan–pectin layer did not inhibit lipase activity, which may have been because the chitosan–pectin desorbed from the droplet surfaces thereby allowing the enzyme to reach the lipids; however, further work is needed to establish this. This information could be used to create food emulsions with low caloric level, or to optimize diets for individuals with lipid digestion problems. 相似文献
15.
16.
Ana Navarro Manuel J. Bández Carmen Gómez Marisa G. Repetto Alberto Boveris 《Journal of bioenergetics and biomembranes》2010,42(5):405-412
Rotenone and pyridaben were tested on activities and properties of rat brain mitochondria determining Ki (inhibitor concentration
at half maximal inhibition) and Imax (% of inhibition at maximal inhibitor concentration). The assayed activities were complexes
I, II and IV, respiration in states 3, 3u (uncoupled) and 4, biochemical and functional activities of mitochondrial nitric
oxide synthase (mtNOS), and inner membrane potential. Selective inhibitions of complex I activity, mitochondrial respiration
and membrane potential with malate-glutamate as substrate were observed, with a Ki of 0.28–0.36 nmol inhibitor/mg of mitochondrial
protein. Functional mtNOS activity was half-inhibited at 0.70–0.74 nmol inhibitor/mg protein in state 3 mitochondria and at
2.52–2.98 nmol inhibitor/mg protein in state 3u mitochondria. This fact is interpreted as an indication of mtNOS being structurally
adjacent to complex I with an intermolecular mtNOS-complex I hydrophobic bonding that is stronger at high Δψ and weaker at
low Δψ. 相似文献
17.
Activities of antioxidant enzymes in mitochondria of growing and dormant sugar beet roots 总被引:1,自引:0,他引:1
A. G. Shugaev D. A. Lashtabega N. A. Shugaeva E. I. Vyskrebentseva 《Russian Journal of Plant Physiology》2011,58(3):387-393
In mitochondria isolated from growing (70–85 days) and dormant (stored for 8–12 weeks) sugar beet (Beta vulgaris L.) roots, activities of superoxide dismutase (SOD) and enzymes of the ascorbate-glutathione cycle were determined. The activity
of SOD, the enzyme involved in superoxide detoxification, was much higher in mitochondria of the growing root, whereas activities
of ascorbate peroxidase (APO) and glutathione reductase (GR), key enzymes of the ascorbate-glutathione cycle involved in the
hydrogen peroxide degradation, increased substantially in mitochondria of dormant storage roots. Catalase (CAT) activity was
detected in the fraction of root mitochondria purified in the sucrose density gradient, which activity was inhibited by cyanide
by 85–90% and much weaker, by aminotriazol (by 30–35%). Submitochondrial localization of APO and CAT was analyzed using proteinase
K. It was established that a substrate-binding APO center is localized on the external side of the inner membrane, whereas
CAT is localized in the mitochondrial matrix. A possible role of mitochondria as ROS (hydrogen peroxide) acceptors in the
cells of storage parenchyma of the stored root is discussed. 相似文献
18.
Tau hyperphosphorylation, amyloid plaques, and neuronal death are major neuropathological features of Alzheimer’s disease
(AD) and Prion-related encephalopathies (PRE). Cyclin-dependent kinase 5 (Cdk5) is a serine/threonine kinase, active in post-mitotic
neurons, where it regulates survival and death pathways. Overactivation of Cdk5 is conferred by p25, a truncated fragment
of the p35 activator formed upon calpain activation. Cdk5 deregulation causes abnormal phosphorylation of microtubule-associated
protein tau, leading to neurodegeneration. In this work we investigated the involvement of Cdk5 in the neurodegeneration triggered
by amyloid-beta (Aβ) and prion (PrP) peptides, the culprit agents of AD and PRE. As a work model, we used cultured rat cortical
neurons treated with Aβ1–40 and PrP106–126 synthetic peptides. The obtained data show that apoptotic neuronal death caused by both the peptides was in part due to Cdk5
deregulation. After peptide treatment, p25 levels were significantly enhanced in a pattern consistent with the augment in
calpain activity. Moreover, Aβ1–40 and PrP106–126 increased the levels of tau protein phosphorylated at Ser202/Thr205. Cdk5 (roscovitine) and calpain (MDL28170) inhibitors
reverted tau hyperphosphorylation and prevented neuronal death caused by Aβ1–40 and PrP106–126. This study demonstrates, for the first time, that Cdk5 is involved in PrP-neurotoxicity. Altogether, our data suggests that
Cdk5 plays an active role in the pathogenesis of AD and PRE. 相似文献
19.
Sławomir Orzechowski Joanna Socha-Hanc Andrzej Paszkowski 《Acta Physiologiae Plantarum》1999,21(4):331-334
The intracellular distribution of alanine aminotransferase (AlaAT, EC 2.6.1.2) activity with L-alanine and 2-oxoglutarate
as a substrates in maize whole leaf extract and bundle sheath cells was studied. After isolation of the mitochondrial-peroxisomal
fraction, mitochondria and peroxisomes were separated by centrifugation on a linear 40–52 % (w/w) sucrose gradient. L-Alanine-2-oxoglutarate
transaminating activity of whole leaf extract showed two peaks: first distinctly higher associated with mitochondria and second
lower with peroxisomes. In bundle sheath cells only one peak of this activity was found. It corresponded to the mitochondrial
region of the gradient. It is proposed that mitochondrial L-alanine — 2-oxoglutarate activity was brought about by AlaAT.
Glycine aminotransferase (EC 2.6.1.4) could be responsible for the same activity in peroxisomes.
This work was supported by the State Committee for Scientific Research, a grant No. 5PO6A00510 相似文献
20.
Nitrogen deposition effects on soil organic matter chemistry are linked to variation in enzymes,ecosystems and size fractions 总被引:2,自引:0,他引:2
A. Stuart Grandy Robert L. Sinsabaugh Jason C. Neff Martina Stursova Donald R. Zak 《Biogeochemistry》2008,91(1):37-49
Recent research has dramatically advanced our understanding of soil organic matter chemistry and the role of N in some organic
matter transformations, but the effects of N deposition on soil C dynamics remain difficult to anticipate. We examined soil
organic matter chemistry and enzyme kinetics in three size fractions (>250 μm, 63–250 μm, and <63 μm) following 6 years of
simulated atmospheric N deposition in two ecosystems with contrasting litter biochemistry (sugar maple, Acer saccharum—basswood, Tilia americana and black oak, Quercus velutina—white oak, Q. alba). Ambient and simulated (80-kg NO3
−–N ha−1 year−1) atmospheric N deposition were studied in three replicate stands in each ecosystem. We found striking, ecosystem-specific
effects of N deposition on soil organic matter chemistry using pyrolysis gas chromatography/mass spectrometry. First, furfural,
the dominant pyrolysis product of polysaccharides, was significantly decreased by simulated N deposition in the sugar maple–basswood
ecosystem (15.9 vs. 5.0%) but was increased by N deposition in the black oak–white oak ecosystem (8.8 vs. 24.0%). Second,
simulated atmospheric N deposition increased the ratio of total lignin derivatives to total polysaccharides in the >250 μm
fraction of the sugar maple–basswood ecosystem from 0.9 to 3.3 but there were no changes in other size classes or in the black
oak–white oak ecosystem. Third, simulated N deposition increased the ratio of lignin derivatives to N-bearing compounds in
the 63–250 and >250 μm fractions in both ecosystems but not in the <63 μm fraction. Relationships between enzyme kinetics
and organic matter chemistry were strongest in the particulate fractions (>63 μm) where there were multiple correlations between
oxidative enzyme activities and concentrations of lignin derivatives and between glycanolytic enzyme activities and concentrations
of carbohydrates. Within silt-clay fractions (<63 μm), these enzyme-substrate correlations were attenuated by interactions
with particle surfaces. Our results demonstrate that variation in enzyme activity resulting from atmospheric N deposition
is directly linked to changes in soil organic matter chemistry, particularly those that occur within coarse soil size fractions.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献