Damage caused to Saccharomyces cerevisiae SY4 plasma membrane H(+)-ATPase by Fe- and Cu-Fenton reagents was determined in secretory vesicles containing enzyme in which Cys residues were replaced singly or in pairs by Ala. Cys-221 situated in a beta-sheet domain between M2 and M3 segments, phosphorylation domain-located Cys-409 and Cys-532 situated at the ATP-binding site play a role in the inactivation. In the presence of all three residues the enzyme exhibited a certain basic inactivation, which did not change when Cys-532 was replaced with Ala. In mutants having intact Cys-532 but lacking one or both other cysteines, replacement of Cys-221 with Ala led to lower inactivation, suggesting that Cys-221 may serve as a target for metal-catalyzed oxidation and intact Cys-532 promotes this target role of Cys-221. In contrast, the absence of Cys-409 caused higher inactivation by Fe-Fenton. Cys-532 thus seems to serve as a target for Fe-Fenton, intact Cys-409 causing a conformational change that makes Cys-532 less accessible to oxidation. The mutant lacking both Cys-221 and Cys-409 is more sensitive to Fe-Fenton than to Cu-Fenton and the absence of both Cys residues thus seems to expose presumable extra Fe-binding sites. These data and those on protection by ATP, ADP, 1,4-dithiothreitol and deferrioxamine B point to complex interactions between individual parts of the enzyme molecule that determine its sensitivity towards Fenton reagents. ATPase fragmentation caused by the two reagents differed in that the Fe-Fenton reagent produced in Western blot "smears" whereas the Cu-Fenton reagent produced defined fragments.

• MeSH
• buněčná membrána účinky léků   enzymologie   MeSH
• cystein metabolismus   MeSH
• měď MeSH
• mutace MeSH
• oxidační stres účinky léků   MeSH
• peroxid vodíku farmakologie   MeSH
• protonové ATPasy chemie   genetika   metabolismus   MeSH
• Saccharomyces cerevisiae chemie   účinky léků   enzymologie   MeSH
• substituce aminokyselin MeSH
• volné radikály MeSH
• vztahy mezi strukturou a aktivitou MeSH
• železo farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• cystein MeSH
• Fenton's reagent MeSH Prohlížeč
• měď MeSH
• peroxid vodíku MeSH
• protonové ATPasy MeSH
• volné radikály MeSH
• železo MeSH

In the facultatively anaerobic yeast Saccharomyces cerevisiae the uptake rate and the accumulation ratio of 2-aminoisobutyric acid was decreased by some 30% by Fenton's reagent (FR), a powerful source of OH. radicals. Likewise, the uptake of glutamic acid, leucine and arginine was diminished. The mediated diffusion of 6-deoxy-D-glucose was not affected. The H+ symport of maltose and trehalose was inhibited by some 40% both in the initial rate and in the accumulation ratio. FR had a dramatic inhibitory effect when present during preincubation with 50 mmol/L glucose. In the obligately aerobic Lodderomyces elongisporus the uptake of all amino acids tested was decreased by 15-30%, that of 6-deoxy-D-glucose by about 10%. The initial rates of uptake of maltose and trehalose were depressed by FR by 40% and the acceleration of uptake observed after 8 min of incubation, was abolished by FR completely. Acidification rate of the external medium by S. cerevisiae in the presence of glucose or galactose was enhanced three-fold, that after subsequently added K+ was substantially decreased. FR appears to have a dual effect on sugar and amino acid transport processes in yeast: (1) it blocks carrier protein synthesis; (2) it inhibits the source of energy for transport. It does not appreciably affect the carrier proteins themselves.

• MeSH
• aktivní transport účinky léků   MeSH
• aminokyseliny metabolismus   MeSH
• fungální proteiny metabolismus   MeSH
• kinetika MeSH
• koncentrace vodíkových iontů MeSH
• metabolismus sacharidů MeSH
• peroxid vodíku farmakologie   MeSH
• Saccharomyces cerevisiae účinky léků   metabolismus   MeSH
• Saccharomycetales účinky léků   metabolismus   MeSH
• transportní proteiny metabolismus   MeSH
• železo farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aminokyseliny MeSH
• Fenton's reagent MeSH Prohlížeč
• fungální proteiny MeSH
• peroxid vodíku MeSH
• transportní proteiny MeSH
• železo MeSH

Oxidative stress in microbial cells shares many similarities with other cell types but it has its specific features which may differ in prokaryotic and eukaryotic cells. We survey here the properties and actions of primary sources of oxidative stress, the role of transition metals in oxidative stress and cell protective machinery of microbial cells, and compare them with analogous features of other cell types. Other features to be compared are the action of Reactive Oxygen Species (ROS) on cell constituents, secondary lipid- or protein-based radicals and other stress products. Repair of oxidative injury by microorganisms and proteolytic removal of irreparable cell constituents are briefly described. Oxidative damage of aerobically growing microbial cells by endogenously formed ROS mostly does not induce changes similar to the aging of multiplying mammalian cells. Rapid growth of bacteria and yeast prevents accumulation of impaired macromolecules which are repaired, diluted or eliminated. During growth some simple fungi, such as yeast or Podospora spp., exhibit aging whose primary cause seems to be fragmentation of the nucleolus or impairment of mitochondrial DNA integrity. Yeast cell aging seems to be accelerated by endogenous oxidative stress. Unlike most growing microbial cells, stationary-phase cells gradually lose their viability because of a continuous oxidative stress, in spite of an increased synthesis of antioxidant enzymes. Unlike in most microorganisms, in plant and animal cells a severe oxidative stress induces a specific programmed death pathway--apoptosis. The scant data on the microbial death mechanisms induced by oxidative stress indicate that in bacteria cell death can result from activation of autolytic enzymes (similarly to the programmed mother-cell death at the end of bacillary sporulation). Yeast and other simple eukaryotes contain components of a proapoptotic pathway which are silent under normal conditions but can be activated by oxidative stress or by manifestation of mammalian death genes, such as bak or bax. Other aspects, such as regulation of oxidative-stress response, role of defense enzymes and their control, acquisition of stress tolerance, stress signaling and its role in stress response, as well as cross-talk between different stress factors, will be the subject of a subsequent review.

• MeSH
• antioxidancia metabolismus   MeSH
• Bacteria cytologie   metabolismus   MeSH
• biologické modely MeSH
• buněčná smrt MeSH
• houby cytologie   metabolismus   MeSH
• kovy metabolismus   MeSH
• metabolismus lipidů MeSH
• mikrobiologie * MeSH
• oxidační stres * MeSH
• poškození DNA MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• rostliny metabolismus   MeSH
• stárnutí buněk MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• srovnávací studie MeSH
• Názvy látek
• antioxidancia MeSH
• kovy MeSH
• reaktivní formy kyslíku MeSH