Yefim Manevich, PhD
Research Associate Professor
Ph.D., M.V. Lomonosov Moscow Institute
(University) of Fine Chemical Technology
Office: DD 410
Oxidative and nitrosative stress effect on cellular physiology with focus on intracellular Ca2+ and NO fluxes and protein thiol redox status modification.
Glutathionylation is a dynamic and functionally important post- translational modification of proteins. The major difference of glutathionylation from other post-translation modifications (glycosilation or phosphorylation) is that glutathionylation/deglutathionylation, in addition to enzyme-mediated catalysis, may occur spontaneously for the protein thiolate anions having basic micro vicinity. Glutathione is a major intracellular component (5-10 mM). The ratio of oxidized (GSSG) to reduced (GSH) glutathione corresponding with cellular redox status. Under normal conditions intracellular redox status is very reductive and its changes correspond to multiple pathological states including cancer. Cellular redox status is directly affected by oxidative/nitrozative stress. The consequences of oxidative/nitrosative stress are site-specific and result in a protein oxidation/nirosylation. In many cases protein thiol are the sites of oxidation/nitrosylation, therefore protein glutathionylation may serve two major purposes: protection of protein-thiol from irreversible oxidation and/or preventing protein misfolding through disulfide formation.
We are focused on studying a general effect of protein glutathionylation on cellular physiology as well as a specific effect of novel anticancer redox-active drug NOV002 and NO-generating drug PABA/NO on cellular plasma membrane function and on intracellular Ca2+ and NO homeostasis. To measure intracellular Ca2+ and NO fluxes, as well as tras-plasma and tras-mitochondrial membrane potentials we are using specific fluorescent probes: Fluo-3, Fura-2, DAF-FM-DA, bisOxanol and JC-9.
Peroxiredoxins are none-Se peroxidases, which control intracellular level of hydrogen peroxide to maintain normal intracellular signaling. There are six isoenzymes in the peroxiredoxin superfamily. The peroxiredoxin VI (Prdx6, 1-cys-Prx) is a unique enzyme, which combines GSH-peroxidase and PLA2 activities specifically towards phospholipid hydroperoxides. Prdx6 activation as GSH-peroxidase depends on its glutathyonylation catalysed by GSTpi. There is several reported genetic variation of GSTpi which association with specific pathological states including cancer is under investigation. Prdx6 inactivation correlates with cellular susceptibility to oxidant stress through specific accumulation of phospholipid hydroperoxide and consequent malfunctioning of biomembrane.
Effect of GSTpi polimorphism on Prdx6 activation in vitro and in tissue culture could be used to predict outcome of an oxidative stress-metiated pathological states including cancer in vivo.
To model membrane-specific effect of radiation we developed chemical equivalents of OH-radical generation and quantification using fluorescent detection with coumarin-3-carboxylic acid. To detect phospholipids peroxidation we are using HPLC, UPLC-MS, DPPP-fluorescent detection in vivo and in tissue culture.
To study protein-protein interaction we are using IP with development of specific affinity mini column, crosslinking with series of biotin-labeled agents, FRET, fluorescence dual-color imaging (Li-CORE), SEC with MS analysis.
Recent Publications | Additional Publications
1. Goetzl L, Manevich Y, Roedner C, Praktish A, Hebbar L, Townsend DM. Maternal and fetal oxidative stress and intrapartum term fever. Am J Obstet Gynecol. 2010 Apr;202(4):363.e1-5. PubMed PMID: 20350644.
2. Manevich Y, Townsend DM, Hutchens S, Tew KD. Diazeniumdiolate mediated nitrosative stress alters nitric oxide homeostasis through intracellular calcium and S-glutathionylation of nitric oxide synthetase. PLoS One. 2010 Nov 30;5(11):e14151. PMID: 21152397
3. Neumann CA, Cao J, Manevich Y. Peroxiredoxin 1 and its role in cell signaling. Cell Cycle. 2009 Dec 15;8(24):4072-8. Epub 2009 Dec 5. Review. PubMed PMID:19923889.
4. Tew KD, Manevich Y, Grek C, Xiong Y, Uys J, Townsend DM. The role of glutathione S-transferase P in signaling pathways and S-glutathionylation in cancer. Free Radic Biol Med. 2011 Jul 15;51(2):299-313. Epub 2011 Apr 22. PMID: 21558000
5. Townsend DM, Manevich Y, He L, Xiong Y, Bowers RR Jr, Hutchens S, Tew KD. Nitrosative stress-induced s-glutathionylation of protein disulfide isomerase leads to activation of the unfolded protein response. Cancer Res. 2009 Oct 1;69(19):7626-34. Epub 2009 Sep 22. PubMed PMID: 19773442; PubMed Central PMCID: PMC2756322.
6. Cao J, Schulte J, Knight A, Leslie NR, Zagozdzon A, Bronson R, Manevich Y, Beeson C, Neumann CA. Prdx1 inhibits tumorigenesis via regulating PTEN/AKT activity. EMBO J. 2009 May 20;28(10):1505-17. Epub 2009 Apr 16. PubMed PMID: 19369943; PubMed Central PMCID: PMC2688529.