Group of Cross-Linking Enzymes

Scientific units

Head: Nikolay Pestov, Ph.D.

GMBS was established de facto in 1979, when Yu.A. Ovchinnikov, vice-president of Academy of Sciences of USSR ordered N.N., Modyanov to organize a new scientific group to pursue a new direction of research primarily aimed at molecular basis of ion transport across biological membranes. On May 19, 1986, the group was given a formal status as a division of Shemyakin Institute of Bioorganic Chemistry - that day may be celebrated as the beginning of its official history. Major efforts of the group were directed towards structure-functional aspects of ion-transporting ATPases, especially of the Na,K-ATPase In colloboration with E.D. Sverdlov, primary structures of α and β-subunits of Na,K-ATPase from pig kidney were determined by DNA sequencing for the first time, independently from and nearly simultaneously with competing groups from USA and Japan, which were studying similar objects from different species. The resulting paper, published in FEBS Letters, was cited by other scientists several hundred times. This success inspired further studies including investigations of Na,K-ATPase transmembrane fold – an absolutely mysterious aspect of molecular organization at that time. For this purpose, various experimental approaches were used, such as limited proteolysis, hydrophobic photoactivatable labeling, vectorial iodination by lactoprexidase, as well as the use of site-specific monoclonal antibodies. A parallel research direction lead to the discovery of novel homologous X,К-ATPase genes, for example, ATP1AL1, later shown to be a novel enzyme, termed nongastric Н,К-ATPase. From 1986 to 1992 the group was one of the leaders among divisions of the Shemyakin Institute as judged by the number of papers in peer-reviewed journals.

A new stage began in 1992 coinciding with unexpected break-up of the USSR and simultaneous departure of of more than half of the group's staff to USA. By now, most of them have built highly successfull careers in science, suffice it to mention Nikolay Modyanov (Toledo University), Svetlana Lutsenko (Oregon Health Science University), Konstantin Petrukhin (Merck Research Laboratories), Alexander Grishin (Mount Sinai School of Medicine), Grigory Belogrudov (University of California, Los Angeles), Elena Arystarkhova (Massachusetts General Hospital and Harvard Medical School).

Remaining collegues had to exert extraordinary efforts in order to prevent the group from disintegration. The leadership was taken by M.I. Shakhparonov, appointed acting head on January 4, 1993, and the official group head from December 30, 1994, whereas the division was officially renamed as Group of Membrane Bioenergetic Systems (November 2, 1994). A deep reorganization was to follow that would help to preserve and raise scientific level by means of acquisition of new equipment and application of modern methods. Thus the group transformed into a highly dynamic, creative and solidary body. The ever persisting dearth of adequate funding was partly alleviated by grants from Russian Foundation for Basic Research, Russian Ministry of Science, Soros International Foundation, INTAS and CRDF.

In recent years, new perspectives have been found and the spectrum of GMBS investigations has been significantly widened thanks to old partnerships and also new ones in Russia and abroad. Especially important, pleasant and fuitful were colloborations with groups of N.N. Modyanov (University of Toledo, USA), K. Geering (University of Lausanne, Switzerland), J. Rydstrom (Goteborg University, Sweden) and E.D. Sverdlov (Shemyakin-Ovchinnikov Institute).

Members of GMBS together with post- and undergraduate students have been studying the diversity of X,К-ATPase isoforms, such as nongastric H,K-ATPase. An important point was the discovery of a new isoform of its β-subunit, termed βm, which possesses some unusual properties. Versatile studies of proton-transporting transhydrogenase, an interesting enzyme of bacteria and mitochondria, constituted another important research field of GMBS. In collaboration with the group of prof. Pepe (Australia) and prof. Ridstrem (Sweden) we have shown the role of transhydrogenase in maintaining antioxidant mitochondrial protection during heart failure.

On November 01, 2009 GMBS was split into two groups: GMBS and Group of Cross-Linking Enzymes (GCLE). GMBS and GKSF continued the established lines of investigation and alo entered several new areas. Main achievements include identification of new protein interactors of lysyl oxidase (LOX), a protein cross-linking enzymes. The most promising protein was p66-β, a component of the nucleosome-remodeling complex Mi2 / NuRD. Studies on the second interesting cross-linking enzyme, transglutaminase TG2, allowed developing a fundamentally new method of detecting its conformational dynamics in living cells. It has been shown that apoptosis activation changes conformation of cytoplasmic TG2 whereas endosomal TG2 remains in a state that lacks the transamidation activity. We succeeded in demonstrating that the interaction of survivin-Ran is extremely important for the resistance glioblastoma, a highly aggressive tumor, to therapy. Disruption of the survivin-Ran complex by a small molecule inhibitor of this protein-protein interaction LLP-3 interferes with the survival and growth of glioblastoma cells both in vitro and in vivo. Further, the phenotype of glioblastoma cells was shown to depend on the proteins that regulate alternative splicing.

Significant differences in the expression of splicing factors between proineuronal and mesenchymal stem cells of glioblastoma have been revealed. It has been demonstrated that protein regulators of splicing play an important role in signaling between tumor cells, and this signaling is carried out by extracellular vesicles.

Nikolay Pestov, Ph.D.depart.
Irina Okkelman, Ph.D.j. r.
Aliya ZakirovaPhD

Former members:

Natalia Gevondyan, Ph.D.s. r.

Selected publications

  1. Dmitriev R.I., Pestov N.B., Shakhparonov M.I., Okkelman I.A. (2014). Two distinct nuclear localization signals in mammalian MSL1 regulate its function. J. Cell. Biochem. 115 (11), 1967–73 [+]

    MSL1 protein regulates global histone H4 acetylation at residue K16 in stem and cancer cells, through interaction with KAT8. The functional significance of mammalian MSL1 isoforms, involved in various protein interactions, is poorly understood. We report the identification of a novel nuclear localization signal (NLS), common to all MSL1 isoforms, in addition to previously known bipartite NLS, located in domain PEHE. Isoforms having both NLS localize to sub-nuclear foci where they can target co-chaperone protein TTC4. However, all MSL1 isoforms also have ability to affect H4K16 acetylation. Thus, presence of two NLS in MSL1 protein can mediate activity of KAT8 in vivo.

  2. Okkelman I.A., Sukaeva A.Z., Kirukhina E.V., Korneenko T.V., Pestov N.B. (2014). Nuclear translocation of lysyl oxidase is promoted by interaction with transcription repressor p66β. Cell Tissue Res. 358 (2), 481–9 [+]

    Lysyl oxidase (LOX) is an amine oxidase involved in protein cross-linking of the extracellular matrix. Less well characterized is the role that LOX plays among nuclear proteins, and molecular mechanisms of its transport to the nucleus are currently unknown. Here, we have employed yeast two-hybrid library screening and found that the LOX catalytic domain interacts with the transcription repressor p66β. This interaction has been confirmed in vitro and has been found to be accomplished through the CR2-containing domain of p66β. Moreover, co-expression of p66β and LOX in living tumor cells leads to the nuclear accumulation of LOX. Thus, p66β might be important for the regulation of LOX in the nucleus.


Nikolay Pestov

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