Effect of cytochrome c on the generation and elimination of O2*- and H2O2 in mitochondria

The primary recognized function of cytochrome c is to act as an electron carrier transferring electrons from complex III to complex IV in the respiratory chain of mitochondria. Recent studies on cell apoptosis reveal that cytochrome c is responsible for the programmed cell death when it is released from mitochondria to cytoplasm. In this study we present evidence showing that cytochrome c plays an antioxidative role by acting on the generation and elimination of O(2)(*) and H(2)O(2) in mitochondria. The O(2)(*) and H(2)O(2) generation in cytochrome c-depleted Keilin-Hartree heart muscle preparation (HMP) is 7-8 times higher than that in normal HMP. The reconstitution of cytochrome c to the cytochrome c-depleted HMP causes the O(2)(*) and H(2)O(2) generation to exponentially decrease. An alternative electron-leak pathway of the respiratory chain is suggested to explain how cytochrome c affects on the generation and elimination of O(2)(*) and H(2)O(2) in mitochondria. Enough cytochrome c in the respiratory chain is needed for keeping O(2)(*) and H(2)O(2) at a lower physiological level. A dramatic increase of O(2)(*) and H(2)O(2) generation occurs when cytochrome c is released from the respiratory chain. The burst of O(2)(*) and H(2)O(2), which happens at the same time as cytochrome c release from the respiratory chain, should have some role in the early stage of cell apoptosis.     

Effects and mechanisms of H(2)O(2) on production of dicarboxylic acid.

The system of producing long chain dicarboxylic acid (DCA) by Candida tropicalis is an aerobic and viscous fermentation system. A method to overcome the gas-liquid transport resistance and to increase oxygen supply is by adding hydrogen peroxide (H(2)O(2)) to the fermentation system. Here we report that the H(2)O(2) not only can enhance the oxygen supply but also change the metabolism by inducing cytochrome P450, the key enzyme of a, o-oxidation. When C. tropicalis was cultivated in a 3-L bioreactor using the combination of aeration and H(2)O(2) feeding, DCA production rates increased by about 10% after a short period of decrease at the beginning. Furthermore, the experiments showed that the maximum activities of P450 could be induced at 2 mM H(2)O(2), and the inducible mechanisms are also discussed. Moreover, we suggest that alkane might be oxidized through the "peroxide shunt pathway" when H(2)O(2) is present. By adding H(2)O(2), the DCA yield in a 22-L bioreactor could increase by 25.3% and reach 153.9 g/L.     

Lag phase of CO2-dependent O2 evolution by illuminated Anabaena variabilis cells.

The steady-state rate of CO2-dependent O2 evolution by Anabaena variabilis cells in response to illumination was established after a lag phase. The lag phase was shortened (1) in cells incubated with glucose as an oxidizable substrate and (2) upon an increase in light intensity. The lag phase was absent during electron transfer from H2O to p-benzoquinone (in combination with ferricyanide) involving Photosystem II. A lag was observed during electron transfer from H2O to methyl viologen involving Photosystems II and I, but not for electron transfer from N,N,N',N'-tetramethyl-p-phenylenediamine (in combination with ascorbate) to methyl viologen involving only Photosystem I. The lag phases of the light-induced H2O --> CO2 and H2O --> methyl viologen electron transfer reactions showed the same temperature dependences at 10-30 degrees C. The lag was prevented by 3-(3,4-dichlorophenyl)-1,1-dimethylurea at concentrations that caused partial inhibition of photosynthetic O2 evolution. Retardation of cell respiration by a combination of CN- and benzylhydroxamate shortened the lag phase of the H2O --> methyl viologen electron transfer. It is concluded that the lag phase is associated with the electron transfer step between Photosystem II and Photosystem I common for the photosynthetic and respiratory chains and is due to the stimulation of cell respiration during the initial period of illumination as a consequence of an increase in the reduced plastoquinone pool and to subsequent retardation of respiration resulting from the transition of the electron transfer chain to the competitive pathway involving Photosystem I.     

cAMP potentiates H(2)O(2)-induced ERK1/2 phosphorylation without the requirement for MEK1/2 phosphorylation

In Jurkat T lymphocytes, hydrogen peroxide (H(2)O(2)) potentiates the phosphorylation level of extracellular signal-regulated kinase 1 and 2 (ERK1/2) caused by T cell receptor (TCR) stimulation with anti-CD3 and anti-CD28 or anti-CD3 alone. Submillimolar concentrations of H(2)O(2)-induced phosphorylation of ERK1/2 and MAP/ERK kinase 1 and 2 (MEK1/2) without antigenic stimulation. H(2)O(2) also induced the electrophoretic mobility shift of Lck from 56 to 60 kDa. The MEK inhibitor, PD98059 attenuated ERK1/2 and MEK1/2 phosphorylation, as well as the migration shift of Lck induced by H(2)O(2). The phospholipase C (PLC) inhibitor, U73122, and EGTA reduced the phosphorylation of both ERK1/2 and MEK1/2 induced by H(2)O(2). Interestingly, an increase of intracellular cAMP level with forskolin or 8-(4-chlorophenylthio)-cAMP augmented ERK1/2 phosphorylation by H(2)O(2), while inhibiting MEK1/2 phosphorylation by H(2)O(2). These results demonstrate an alternative pathway that results in augmentation of ERK1/2 phosphorylation without concomitant MEK1/2 phosphorylation in T cells.     

ATP对中性粒细胞H2O2产生双重作用机制的研究

本文通过高压液相法测定ATP的代谢,探讨其对中性粒细胞H2O2产生双重作用的机制。结果显示,ATP本身不能激活中性粒细胞产生H     

Dopamine-derived dopaminochrome promotes H(2)O(2) release at mitochondrial complex I: stimulation by rotenone, control by Ca(2+), and relevance to Parkinson disease

Inhibitors of Complex I of the mitochondrial respiratory chain, such as rotenone, promote Parkinson disease-like symptoms and signs of oxidative stress. Dopamine (DA) oxidation products may be implicated in such a process. We show here that the o-quinone dopaminochrome (DACHR), a relatively stable DA oxidation product, promotes concentration (0.1-0.2 mum)- and respiration-dependent generation of H(2)O(2) at Complex I in brain mitochondria, with further stimulation by low concentrations of rotenone (5-30 nm). The rotenone effect required that contaminating Ca(2+) (8-10 mum) was not removed. DACHR apparently extracts an electron from the constitutively autoxidizable site in Complex I, producing a semiquinone, which then transfers an electron to O(2), generating O(2)(.) and then H(2)O(2). Mitochondrial removal of H(2)O(2) monoamine, formed by either oxidase activity or DACHR, was performed largely by glutathione peroxidase and glutathione reductase, which were negatively regulated by low intramitochondrial Ca(2+) levels. Thus, the H(2)O(2) formed accumulated in the medium if contaminating Ca(2+) was present; in the absence of Ca(2+), H(2)O(2) was completely removed if it originated from monoamine oxidase, but was less completely removed if it originated from DACHR. We propose that the primary action of rotenone is to promote extracellular O(2)(.) release via activation of NADPH oxidase in the microglia. In turn, O(2)(.) oxidizes DA to DACHR extracellularly. (The reaction is favored by the lack of GSH, which would otherwise preferably produce GSH adducts of dopaminoquinone.) Once formed, DACHR (which is resistant to GSH) enters neurons to activate the rotenone-stimulated redox cycle described.     

Regulation of hydrogen peroxide production by brain mitochondria by calcium and Bax

Abnormal accumulation of Ca2+ and exposure to pro-apoptotic proteins, such as Bax, is believed to stimulate mitochondrial generation of reactive oxygen species (ROS) and contribute to neural cell death during acute ischemic and traumatic brain injury, and in neurodegenerative diseases, e.g. Parkinson's disease. However, the mechanism by which Ca2+ or apoptotic proteins stimulate mitochondrial ROS production is unclear. We used a sensitive fluorescent probe to compare the effects of Ca2+ on H2O2 emission by isolated rat brain mitochondria in the presence of physiological concentrations of ATP and Mg2+ and different respiratory substrates. In the absence of respiratory chain inhibitors, Ca2+ suppressed H2O2 generation and reduced the membrane potential of mitochondria oxidizing succinate, or glutamate plus malate. In the presence of the respiratory chain Complex I inhibitor rotenone, accumulation of Ca2+ stimulated H2O2 production by mitochondria oxidizing succinate, and this stimulation was associated with release of mitochondrial cytochrome c. In the presence of glutamate plus malate, or succinate, cytochrome c release and H2O2 formation were stimulated by human recombinant full-length Bax in the presence of a BH3 cell death domain peptide. These results indicate that in the presence of ATP and Mg2+, Ca2+ accumulation either inhibits or stimulates mitochondrial H2O2 production, depending on the respiratory substrate and the effect of Ca2+ on the mitochondrial membrane potential. Bax plus a BH3 domain peptide stimulate H2O2 production by brain mitochondria due to release of cytochrome c and this stimulation is insensitive to changes in membrane potential.     

Roles of RhoA and phospholipase A2 in the elevation of intracellular H2O2 by transforming growth factor-beta in Swiss 3T3 fibroblasts.

We have investigated the mechanisms by which transforming growth factor-beta (TGF-beta) increased intracellular H2O2 in Swiss 3T3 fibroblasts. Increase of intracellular H2O2 by TGF-beta was maximal at 30 min and blocked by catalase from Aspergillus niger. Scrape-loading of C3 transferase, which down-regulated RhoA, inhibited the production of H2O2 in response to TGF-beta. TGF-beta stimulated release of arachidonic acid, which was completely inhibited by mepacrine, a phospholipase A2 inhibitor. Mepacrine also blocked the increase of H2O2 by TGF-beta. In addition, arachidonic acid increased intracellular H2O2. Furthermore, TGF-beta stimulated stress fibre formation, which was blocked by catalase, without membrane ruffling. Catalase also inhibited stimulation of thymidine incorporation by TGF-beta. These results suggested that TGF-beta increased intracellular H2O2 through RhoA and phospholipase A2, and also suggested that intracellular H2O2 was required for the stimulation of stress fibre formation and DNA synthesis in response to TGF-beta.     

Peroxiredoxin II functions as a signal terminator for H2O2-activated phospholipase D1

Phospholipase D1 (PLD1) is a signal-transduction regulated enzyme which regulates several cell intrinsic processes including activation of NAPDH oxidase, which elevates intracellular H2O2. Several proteins have been reported to interact with PLD1 in resting cells. We sought to identify proteins that interact with PLD1 after phorbol 12-myristate 13-acetate (PMA) stimulation. A novel interaction with peroxiredoxin II (PrxII), an enzyme that eliminates cellular H2O2, which is a known stimulator of PLD1, was identified by PLD1-affinity pull-down and MS. PMA stimulation was confirmed to promote physical interaction between PLD1 and PrxII and to cause PLD1 and PrxII to colocalize subcellularly. Functional significance of the interaction was suggested by the observation that over-expression of PrxII specifically reduces the response of PLD1 to stimulation by H2O2. These results indicate that PrxII may have a signal-terminating role for PLD1 by being recruited to sites containing activated PLD1 after cellular stimulation involving production of H2O2.     

Chemiluminescence detection of H2O2 produced by human neutrophils during the respiratory burst

A sensitive luminol-dependent chemiluminescence assay for H2O2 was developed for the indirect determination of the transient changes in NADPH oxidase activity associated with the respiratory burst of human neutrophils. A relatively large, controlled amount of horseradish peroxidase was used in combination with added luminol to rapidly remove and simultaneously detect H2O2 as soon as it is formed, thus preventing its accumulation during burst activity and minimizing the effects of side reactions. Cell-derived myeloperoxidase and possibly catalase were inhibited with 90 microM sodium azide to maintain the total catalytic activity toward H2O2 at a constant level. Chemiluminescence measurements of the respiratory burst activity of human neutrophils stimulated with N-formyl-Met-Leu-Phe (fMLP) were in good agreement with measurements made using an established fluorometric assay based on similar principles (P. A. Hyslop and L. A. Sklar (1984) Anal. Biochem. 141, 280-286). In contrast to fluorometry, the chemiluminescence progress curves reflect the instantaneous rather than the integrated levels of H2O2 at any time and are thus a more direct measure of the activity of the NADPH oxidase. This advantage, as well as higher signal-to-noise ratios and greater inherent sensitivity, distinguishes chemiluminescence as a means of following burst activity. The onset of fMLP-stimulated H2O2 generation was detectable by chemiluminescence within 2 s of stimulation (as opposed to more than double this time by fluorometry), showing that high sensitivity is an important consideration in evaluating respiratory burst kinetics. In contrast to fMLP stimulation, longer and concentration-dependent onset times were observed when phorbol myristate acetate was used as a stimulus.     

Increase in External Glutamate and NMDA Receptor Activation Contribute to H2O2-Induced Neuronal Apoptosis

The present study aims to investigate the role of extracellular glutamate and NMDA receptor stimulation in the neuronal death induced by a transient exposure to H2O2 of cultured neurons originating from mouse cerebral cortex. Most of the neuronal loss following a transient exposure to H2O2 of cortical neurons results from an apoptotic process involving a secondary stimulation of NMDA receptors, which occurs after H2O2 washout. Indeed, (a) the neurotoxic effect of H2O2 was strongly reduced by antagonists of NMDA receptors, (b) the neurotoxic effect of H2O2 was enhanced in the absence of Mg2+, (c) the protective effect of MK-801 progressively decayed when it was applied with increasing delay time after H2O2 exposure, and (d), finally, the extracellular concentration of glutamate was increased after H2O2 exposure. The major part of H2O2-induced neurotoxicity is mediated by the formation of hydroxyl radicals, which might be involved in (a) the delayed accumulation of extracellular glutamate and NMDA receptor activation and (b) the poly(ADP-ribose) polymerase activation and the related NAD content decrease. The combination of these two mechanisms could lead to both an increase in ATP consumption and a decrease of ATP synthesis. The resulting large decrease in ATP content might be finally responsible for the neuronal death.     

The nitrone spin trap PBN alters the cellular response to H(2)O(2): activation of the EGF receptor/ERK pathway

The nitrone spin trap PBN has been shown to protect neuronal cells from reactive oxygen species both in culture and in vivo. As an approach to understanding the molecular mechanisms by which PBN may function to protect cells, we examined whether PBN alters the cellular response to reactive oxygen species. H(2)O(2) stimulation of PC-12 cells results in weak activation of both the ERK and JNK signal transduction pathways. PBN pretreatment of PC-12 cells, followed by H(2)O(2) stimulation, results in strong and selective activation of the pro-survival ERK pathway. H(2)O(2) induction of ERK activity in PBN-pretreated cells was shown to be dependent on extracellular Ca(+2) influx. Further analysis of the ERK pathway showed that in PBN-pretreated cells, EGF receptor and the adapter protein SHC were phosphorylated in a Ca(+2)-dependent, ligand-independent manner following H(2)O(2) stimulation. Interestingly, H(2)O(2) stimulation of PBN-pretreated cells results in only 30% of the increase in intracellular Ca(+2) as compared to untreated cells following H(2)O(2) stimulation. These data suggest a model in which PBN attenuates H(2)O(2)-induced Ca(+2) entry, yet magnifies or alters Ca(+2) action, resulting in the activation of the EGF receptor/ERK pathway.     

Effect of voluntary exercise on H2O2 release by subsarcolemmal and intermyofibrillar mitochondria

Previous data have demonstrated that, to handle the oxidative stress encountered with training at high intensity, skeletal muscle relies on an increase in mitochondrial biogenesis, a reduced H(2)O(2) production, and an enhancement of antioxidant enzymes. In the present study, we evaluated the influence of voluntary running on mitochondrial O(2) consumption and H(2)O(2) production by intermyofibrillar mitochondria (IFM) and subsarcolemmal mitochondria (SSM) isolated from oxidative muscles in conjunction with the determination of antioxidant capacities. When mitochondria are incubated with succinate as substrate, both maximal (state 3) and resting (state 4) O(2) consumption were significantly lower in SSM than in IFM populations. Mitochondrial H(2)O(2) release per unit of O(2) consumed was 2-fold higher in SSM than in IFM. Inhibition of H(2)O(2) formation by rotenone suggests that complex I of the electron transport chain is likely the major physiological H(2)O(2)-generating system. In Lou/C rats (an inbred strain of rats of Wistar origin), neither O(2) consumption nor H(2)O(2) release by IFM and SSM were affected by long-term, voluntary wheel training. In contrast, glutathione peroxidase and catalase activity were significantly increased despite no change in oxidative capacities with long-term, voluntary exercise. Furthermore, chronic exercise enhanced heat shock protein 72 accumulation within skeletal muscle. It is concluded that the antioxidant status of muscle can be significantly improved by prolonged wheel exercise without necessitating an increase in mitochondrial oxidative capacities.     

Bovine aortic endothelial cells release hydrogen peroxide.

Endothelial cells grown on microcarriers are able to release H2O2 to the extracellular environment without any added stimulus. The extracellularly released H2O2 can be detected by luminol-amplified chemiluminescence (CL) if horseradish peroxidase is added. The CL response can be reduced by catalase and blocked by superoxide dismutase, indicating that O2- could be a precursor for H2O2. The CL kinetics, i.e., a long lag time followed by a rapid shift to a new level, indicate activation of an O2(-)-producing enzyme. The cells are also able to protect themselves from H2O2 stimulation by both catalase and the glutatione system. Bradykinin stimulates the H2O2 release, but if the effect is directly stimulatory or if it acts by reduction of the protective system is at present unclear. The extracellularly released H2O2 could be a cause of injury to the endothelial cells or to the subendothelial matrix.     

H2O2 generation is decreased by calcium in isolated brain mitochondria

Release of H(2)O(2) in response to Ca(2+) loads (1-100 microM) was investigated using Amplex red fluorescent assay in isolated guinea-pig brain mitochondria respiring on glutamate plus malate or succinate. In mitochondria challenged with Ca(2+) (10 microM), in the absence of adenine nucleotides and inhibitors of the respiratory chain, the rate of H(2)O(2) release, taken as an indication of H(2)O(2) production, was decreased by 21.8+/-1.6% in the presence of NADH-linked substrates and by 86.5+/-1.8% with succinate. Parallel with this, a Ca(2+)-induced loss in NAD(P)H fluorescence, sustained depolarization, decrease in fluorescent light scattering signal and in calcein fluorescence were detected indicating an increased permeability and swelling of mitochondria, which were prevented by ADP (2 mM). In the presence of ADP H(2)O(2) release from mitochondria was decreased, but Ca(2+) no longer influenced the generation of H(2)O(2). We suggest that the decreased H(2)O(2) generation induced by Ca(2+) is related to depolarization and NAD(P)H loss resulting from a non-specific permeability increase of the mitochondrial inner membrane.     

Phosphorylation by Protein Kinase C of Annexin 2 in Chromaffin Cells Stimulated by Nicotine

Abstract: Annexin 2 phosphorylated in vitro by protein kinase C has been shown to restore partially catecholamine secretion in streptolysin O-permeabilized chromaffin cells depleted of their protein kinase C activity. This result suggested a phosphorylation of annexin 2 in stimulated cells. Nicotine stimulation induced an increase of ³²P incorporation in annexin 2 heavy chain concomitant with catecholamine release. This incorporation results from phosphorylation by protein kinase C because (a) serine was the only phosphorylated residue, (b) ³²P incorporation was inhibited by the protein kinase inhibitors H7, GF 109203X, and staurosporine, and (c) activators of this enzyme, 12- O -tetradecanoylphorbol 13-acetate and 1,2-dioctanoylglycerate, increased the incorporation of radioactivity. The phosphorylated heavy chain had an electrophoretic mobility lower than that of the unmodified one, thus allowing determination of the fraction of phosphorylated protein. In the resting state, a significant fraction of annexin 2 heavy chain was phosphorylated, and nicotine stimulation resulted in an activation of both phosphorylation and dephosphorylation. Phosphorylation was largely increased in the presence of okadaic acid, indicating the involvement of type 1 and 2A phosphatases.     

过氧化氢胁迫促进产朊假丝酵母合成谷胱甘肽

谷胱甘肽(GSH)在生物细胞抵御外界环境条件的刺激和胁迫时起到非常重要的作用。考察了不同时间不同浓度过氧化氢胁迫和过氧化氢连续胁迫对产朊假丝酵母合成GSH的影响, 发现低浓度过氧化氢的连续胁迫对GSH的合成有明显促进作用。进一步在发酵罐上应用了低浓度过氧化氢(36 mmol/L)持续胁迫策略, 最终GSH产量为922 mg/L, 胞内GSH含量为1.64%, 比对照分别提高了7%和35%。     

Phosphorylation of proteins and apoptosis induced by c-Jun N-terminal kinase1 activation in rat cardiomyocytes by H(2)O(2) stimulation

Cytokines and various cellular stresses are known to activate c-Jun N-terminal kinase-1 (JNK1), which is involved in physiological function. Here, we investigate the activation of JNK1 by oxidative stress in H9c2 cells derived from rat cardiomyocytes. H(2)O(2) (100 microM) significantly induces the tyrosine phosphorylation of JNK1 with a peak 25 min after the stimulation. The amount of JNK1 protein remains almost constant during stimulation. Immunocytochemical observation shows that JNK1 staining in the nucleus is enhanced after H(2)O(2) stimulation. To clarify the physiological role of JNK1 activation under these conditions, we transfected antisense JNK1 DNA into H9c2 cells. The antisense DNA (2 microM) inhibits JNK1 expression by 80% as compared with expression in the presence of the sense DNA, and significantly blocks H(2)O(2)-induced cell death. Consistent with the decrease in cell number, we detected condensation of the nuclei, a hallmark of apoptosis, 3 h after H(2)O(2) stimulation in the presence of the sense DNA for JNK1. The antisense DNA of JNK1 inhibits the condensation of nuclei by H(2)O(2). Under these conditions, the H(2)O(2)-induced phosphorylation of proteins with molecular masses of 55, 72, and 78 kDa is blocked by treatment with the antisense DNA for JNK1 as compared with the sense DNA for JNK1. These findings suggest that JNK1 induces apoptotic cell death in response to H(2)O(2), and that the cell death may be involved in the phosphorylations of 55, 72, and 78 kDa proteins induced by JNK1 activation.     

Stimulation of respiratory burst by cyclocommunin in rat neutrophils is associated with the increase in cellular Ca2+ and protein kinase C activity

In this study, the underlying mechanisms of stimulation by cyclocommunin, a natural pyranoflavonoid, of respiratory burst in rat neutrophils was investigated. Cyclocommunin evoked a concentration-dependent stimulation of superoxide anion (O2*-) generation with a slow onset and long lasting profile. The maximum response (16.4+/-2.3 nmol O2*-/10 min per 10(6) cells) was observed at 3-10 microM cyclocommunin. Cyclocommunin did not activate NADPH oxidase in a cell-free system. Cells pretreated with pertussis toxin or n-butanol did not affect the cyclocommunin-induced O2*- generation. However, a protein kinase inhibitor staurosporine and EGTA greatly reduced the O2*-generation caused by cyclocommunin. Treatment of neutrophils with phorbol 12-myristate 13-acetate (PMA), but not with formylmethionyl-leucyl-phenylalanine (fMLP), for 20 min significantly reduced the O2*- generation following the subsequent stimulation of cells with cyclocommunin. Cyclocommunin did not affect the cellular mass of phosphatidic acid (PA). Neither the tyrosine kinase inhibitor, genistein, nor the p38 mitogen-activated protein kinase (MAPK) inhibitor, SB203580, affected cyclocommunin-induced O2*- generation. The enzyme activities of neutrophil cytosolic and membrane-associated protein kinase C (PKC) were both increased significantly with 100 microM cyclocommunin. The membrane-associated PKC-theta and PKC-beta were increased following the stimulation of neutrophils with 30 and 100 microM cyclocommunin, respectively. Cyclocommunin reduced the [3H]phorbol 12,13-dibutyrate ([3H]PDB) binding to cytosolic PKC in a concentration-dependent manner. Cyclocommunin (> or =3 microM) significantly evoked a slow and long lasting [Ca2+]i elevation in neutrophils, and a phospholipase C (PLC) inhibitor U73122 greatly inhibited these Ca2+ responses. Moreover, the increase in cellular inositol bis- and trisphosphate (IP2 and IP3) levels were observed in neutrophils stimulated with 30 microM cyclocommunin for 3 min. Collectively, these results indicate that the stimulation of respiratory burst by cyclocommunin is probably mediated by the synergism of PKC activation and [Ca2+]i elevation in rat neutrophils.     

过氧化氢胁迫促进产朊假丝酵母合成谷胱甘肽

谷胱甘肽(GSH)在生物细胞抵御外界环境条件的刺激和胁迫时起到非常重要的作用。考察了不同时间不同浓度过氧化氢胁迫和过氧化氢连续胁迫对产朊假丝酵母合成GSH的影响, 发现低浓度过氧化氢的连续胁迫对GSH的合成有明显促进作用。进一步在发酵罐上应用了低浓度过氧化氢(36 mmol/L)持续胁迫策略, 最终GSH产量为922 mg/L, 胞内GSH含量为1.64%, 比对照分别提高了7%和35%。