LABORATORY OF NUCLEIC ACID-PROTEIN INTERACTIONS
(NAPI Laboratory)

Yu.F.Drygin
Yuri F. Drygin
 Head of Laboratory, Dr.Sci., Ph.D.

e-mail: drygin@belozersky.msu.ru Tel.:(+7 - 495)939-55-29; Fax: (+7-495)939-31-81

Area of expertise:
Chemistry and Biochemistry of Nucleic Acids and Nucleoproteins. Molecular Virology. Molecular Diagnostics of Plant and Animal Virus Infections. Electron and Atomic Force Microscopy of Nucleic Acids and Nucleoproteins. Spiral Plant Virus as Nano Container and Vehicle for Addressed Delivery of Biologically Active Substances in Biology and Medicine.

Team 2010
           Olga A. Kondakova, Ph.D.;
           Anatoly N. Blintzov, Ph.D.
           Elena S. Gavriushina, Ph.D. student,
           Anna D. Sushko, Ph.D. student,
           Anna N. Starovoytova - 3d-year education student

 

Research:

Study of Natural Covalent Complexes of Nucleic Acids and Proteins
.
The main topic of research of the laboratory focuses on isolation, structural and functional study of virus and cell covalent complexes of nucleic acids and proteins. The data obtained are reviewed in the survey:


Yu.F.Drygin "Natural covalent complexes of nucleic acids and proteins: some comments on practice and theory on the path from well-known complexes to new ones" Nucl. Acids Res. (1998), v.26, N21, 4791-4796.

High sensitive tools and methods are developed in the laboratory for detection and identification of the viral and cellular nucleic acids and covalent complexes of RNA and proteins that contain the covalent linkage unit (CLU), which is identical to the CLU of the picornavirus VPg-RNA covalent complex.

Mechanisms of «Formation-Dissociation» of the complexes are also under consideration. Several papers are published on the substrate specificity of the VPg-unlinking enzyme that  catalyzes hydrolysis of the phosphodiester bond between picornavirus RNA and VPg
:

Antibodies against picornavirus RNA-VPg covalent linkage unit
Several model compounds mimicking the structure of the covalent linkage unit of the RNA-VPg complex of picornaviruses were synthesized.

A.A. Shabanov, A.A. Kaluzhny, M.B. Gottikh, Yu.F.Drygin "Natural substrates and its synthetic models of the uridilylpolynucleotide-(5'P'O)-tyrosine phosphodiesterase - an enzyme which hydrolyzes a covalent bond between RNA and VPg of picornaviruses", Biokhimia, v. 61, N6, 1106 -1118 (1996).
O.A. Ivanova, A.G. Venyaminova, M.N. Repkova, Yu.F. Drygin "Polyclonal antibodies against a structure mimicking the covalent linkage unit between picornavirus RNA and VPg: an immunochemical study" Biochemistry (Moscow), 70, 1258 - 1266 (2005).


In 2009/10 we developed efficient synthesis in solution of the model compound, [N(Ac),CO(NHMe)]Tyr-(5'P?O)Up-O-(CH2)6NH2, containing CLU (Tyr-pUp) of the picornavirus VPg-RNA covalent complex. We obtained polyclonal antibodies in rabbit to the hapten which recognized picornavirus covalent VPg-RNA complexes and discovered by immunofluorescent microscopy that staining of the HRV2 (picornavirus) infected cells with antibodies to the CLU and HRV2 VPg (kindly provided by Dr. D.Blaas) are overlapped and localized in sites of the picornavirus RNA replication (Ph.D. Thesis "Antibodies to the covalent linkage unit of the virus RNA-VPg covalent complex" by E.S. Gavruishina).

We propose to use the CLU antibodies as a tool for identification as in vitro so in vivo of the virus and hypothetic cellular covalent complexes of RNA and proteins containing the CLU structure.

Atomic Force microscopy of RNA and ribonucleoproteins
We use AFM and TEM to study viruses, ribonucleoproteins and single-stranded RNA. Joint AFM and electron microscopy study with Dr. D. Blaas (Max F. Perutz Laboratories, Medical University of Vienna) on the mechanism of RNA exit from HRV2 and Mengo virions is in the progress.

E.V. Dubrovin, Yu.F. Drygin, V.K. Novikov, I.V. Yaminsky "Atomic force microscopy as a tool of inspection of viral infection" Nanomedicine: Nanotechnology, Biology, and Medicine" v.3, N 2, 128-131 (2007).
A.D.Sushko, I.V. Yaminsky, E.S. Gavriushina, Yu.F. Drygin "Exit RNA from HRV2 virion in acidified medium", Colloid J., (2010), 72, N 3, p. 1-6.

Molecular Diagnostics of Virus Infections
Combined technology of DNA probe-ELISA, RT-PCR-DNA probe-ELISA, DNA-microplate (DNA array) technologies, and immunostrip chromatography on test-strips were developed by us for laboratory detection of viruses and virus RNAs and field diagnostics of the potato virus and viroid infections.

Yu.F. Drygin, S.N. Chirkov, O.A. Kondakova, R.A. Zinovkin, P.A. Ivanov, A.N. Blintsov, E.S. Gavryushina, A.V. Zherdev, N.A. Byzova, B. B. Dzantiev, J.G. Atabekov "High-sensitive technologies for molecular diagnostics of potato virus and viroid infections". Potato Production and Innovative Technologies (eds. A.J. Haverkort, B.V. Anisimov), Wageningen Acad.Publishers, 2007, pp.274-285.
Yu.F. Drygin, A. N. Blintsov, A. P. Osipov, V. G. Grigorenko, I. P. Andreeva, A. I. Uskov, Yu. A. Varitsev, B. V. Anisimov, V. K. Novikov, J. G. Atabekov "High-sensitivity express immunochromatographic method for detection of plant infection by tobacco mosaic virus" Biochemistry Moscow , 74, 986-983 (2009)

During the course of this work, methods of RNA isolation and detection from bacteria, plants and animal cells with water-soluble non-toxic agent were developed and patented.

Yu.F. Drygin, O.A. Kondakova, S.N. Chirkov, E.S. Gavryushina, J.G. Atabekov (WO/2008/150187) "USE OF AMMONIUM TRICHLOROACETATE IN THE FORM OF AN AGENT FOR DISSOCIATING NATURAL NUCLEIC ACID COMPLEXES AND A RNA EXTRACTING METHOD"

DRYGIN,Yu.F.. ATABEKOV, J.G., KONDAKOVA,O.A.,CHIRKOV,S.N., ZINOVKIN,R.A., KISELEVA,V.I. (WO/2009/123494) "METHOD FOR SIMULTANEOUSLY DETECTING A PLURALITY OF RNA SEQUENCES IN A BIOLOGICAL SAMPLE".

Perspectives: antibodies against virus genomes

In the course of the research we discovered that antisera raised against virus RNAs contain antibodies that reveal specificity to the selective antigenic epitopes (ribotopes) and recognize RNA antigens variously. We are looking at students for this work.

Open Positions
NAPI Laboratory offers positions in the area of molecular biology of nucleoproteins, virology and nanomedicine for excellent and highly motivated biologists and chemists at under-graduate or post-graduate level.

Students: We are constantly offering diploma thesis and PhD thesis projects for students in molecular biology of the nucleoproteins, virology and molecular diagnostics of virus infections. Work in nanomedicine is in progress. The applicant is expected to have a strong interest in experimental activity.



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