Research
The primary focus of my laboratory is studies of protein structure, dynamics and interactions using structural biology methods, including nuclear magnetic resonance (NMR) spectroscopy and X-ray crystallography. We make use of cutting-edge TROSY-NMR techniques that allow to access structural dynamics and interactions of protein assemblies with molecular weights of up to 1 MDa, opening an avenue for deciphering molecular mechanisms of their action.
DNA damage tolerance and cancer:
The underlying cause of cancer is spontaneous mutations introduced to genomic DNA. Reactive products of cellular metabolism and external genotoxic agents cause persistent DNA damage, which is constantly removed through various DNA repair mechanisms. It is unavoidable, however, that some DNA modifications (lesions) persist into S-phase, creating blocks for progression of the DNA replication machinery. To circumvent this problem organisms in all kingdoms of life have evolved DNA damage tolerance pathways, employing specialized enzymes that bypass DNA lesions while temporarily leaving DNA damage unrepaired. The vast majority of mutations are introduced in the genome by enzymes of error-prone branch of DNA damage tolerance - translesion DNA synthesis (TLS). Genetic changes that ensue as a result of TLS are at the root of the onset of cancer and the development of various resistance mechanisms displayed by relapsed tumors, which represents a major problem for treatment of some types of cancer, including ovarian and lung. Our research is aimed at obtaining a detailed atomic-resolution picture of structure, dynamics and interactions of proteins and protein assemblies involved in DNA damage tolerance pathways that will aid the development of new strategies for cancer therapy.
Protein folding and dynamics:
Intermediate and transition states of biomolecular processes represent a paradigm of functionally important structure in biology. For example, protein self-assembly involves the formation of partially folded and misfolded protein states prone to aggregation implicated in a number of human disorders, including type-II diabetes, Alzheimer's and Parkinson's diseases. Although the characterization of such species can provide vital clues about the mechanisms of the underlying processes, it is extremely challenging to examine such states because they are populated at low levels and are not readily isolated. One of the research directions in my laboratory is studies of intermediate and transition states of protein folding and binding using novel NMR relaxation dispersion methodology.
REV1 C-terminal Domain
SHPRH PHD Domain
REV1 BRCT Domain
Rad18 UBZ Domain in Complex with Ubiquitin
Publications
1: Zhuravleva A, Korzhnev DM. Protein folding by NMR. Prog Nucl Magn Reson
Spectrosc. 2017 May;100:52-77. doi: 10.1016/j.pnmrs.2016.10.002. Epub 2016 Nov 9.
Review. PubMed PMID: 28552172.
2: Sail V, Rizzo AA, Chatterjee N, Dash RC, Ozen Z, Walker GC, Korzhnev DM,
Hadden MK. Identification of Small Molecule Translesion Synthesis Inhibitors That
Target the Rev1-CT/RIR Protein-Protein Interaction. ACS Chem Biol. 2017 Jul
21;12(7):1903-1912. doi: 10.1021/acschembio.6b01144. Epub 2017 Jun 9. PubMed
PMID: 28541665.
3: Kukic P, Pustovalova Y, Camilloni C, Gianni S, Korzhnev DM, Vendruscolo M.
Structural Characterization of the Early Events in the Nucleation-Condensation
Mechanism in a Protein Folding Process. J Am Chem Soc. 2017 May
24;139(20):6899-6910. doi: 10.1021/jacs.7b01540. Epub 2017 May 10. PubMed PMID:
28401755.
4: Alyami EM, Rizzo AA, Beuning PJ, Korzhnev DM. NMR resonance assignments for
the N-terminal domain of the δ subunit of the E. coli γ clamp loader complex.
Biomol NMR Assign. 2017 Mar 6. doi: 10.1007/s12104-017-9741-z. [Epub ahead of
print] PubMed PMID: 28265855.
5: Korzhnev DM, Neculai D, Dhe-Paganon S, Arrowsmith CH, Bezsonova I. Solution
NMR structure of the HLTF HIRAN domain: a conserved module in SWI2/SNF2 DNA
damage tolerance proteins. J Biomol NMR. 2016 Nov;66(3):209-219. Epub 2016 Oct
22. PubMed PMID: 27771863.
6: Korzhnev DM, Hadden MK. Targeting the Translesion Synthesis Pathway for the
Development of Anti-Cancer Chemotherapeutics. J Med Chem. 2016 Oct
27;59(20):9321-9336. Epub 2016 Jul 19. Review. PubMed PMID: 27362876.
7: Pustovalova Y, Magalhães MT, D'Souza S, Rizzo AA, Korza G, Walker GC, Korzhnev
DM. Interaction between the Rev1 C-Terminal Domain and the PolD3 Subunit of Polζ
Suggests a Mechanism of Polymerase Exchange upon Rev1/Polζ-Dependent Translesion
Synthesis. Biochemistry. 2016 Apr 5;55(13):2043-53. doi:
10.1021/acs.biochem.5b01282. Epub 2016 Mar 24. PubMed PMID: 26982350; PubMed
Central PMCID: PMC4898654.
8: Pozhidaeva AK, Mohni KN, Dhe-Paganon S, Arrowsmith CH, Weller SK, Korzhnev DM,
Bezsonova I. Structural Characterization of Interaction between Human
Ubiquitin-specific Protease 7 and Immediate-Early Protein ICP0 of Herpes Simplex
Virus-1. J Biol Chem. 2015 Sep 18;290(38):22907-18. doi: 10.1074/jbc.M115.664805.
Epub 2015 Jul 29. PubMed PMID: 26224631; PubMed Central PMCID: PMC4645603.
9: Pustovalova Y, Kukic P, Vendruscolo M, Korzhnev DM. Probing the Residual
Structure of the Low Populated Denatured State of ADA2h under Folding Conditions
by Relaxation Dispersion Nuclear Magnetic Resonance Spectroscopy. Biochemistry.
2015 Aug 4;54(30):4611-22. doi: 10.1021/acs.biochem.5b00345. Epub 2015 Jul 21.
PubMed PMID: 26115097.
10: Kile AC, Chavez DA, Bacal J, Eldirany S, Korzhnev DM, Bezsonova I, Eichman
BF, Cimprich KA. HLTF's Ancient HIRAN Domain Binds 3' DNA Ends to Drive
Replication Fork Reversal. Mol Cell. 2015 Jun 18;58(6):1090-100. doi:
10.1016/j.molcel.2015.05.013. Epub 2015 Jun 4. PubMed PMID: 26051180; PubMed
Central PMCID: PMC4475461.
11: Rizzo AA, Salerno PE, Bezsonova I, Korzhnev DM. NMR structure of the human
Rad18 zinc finger in complex with ubiquitin defines a class of UBZ domains in
proteins linked to the DNA damage response. Biochemistry. 2014 Sep
23;53(37):5895-906. doi: 10.1021/bi500823h. Epub 2014 Sep 15. PubMed PMID:
25162118.
12: Machado LE, Pustovalova Y, Kile AC, Pozhidaeva A, Cimprich KA, Almeida FC,
Bezsonova I, Korzhnev DM. PHD domain from human SHPRH. J Biomol NMR. 2013
Aug;56(4):393-9. doi: 10.1007/s10858-013-9758-2. Epub 2013 Aug 2. PubMed PMID:
23907177; PubMed Central PMCID: PMC3905319.
13: Pustovalova Y, Maciejewski MW, Korzhnev DM. NMR mapping of PCNA interaction
with translesion synthesis DNA polymerase Rev1 mediated by Rev1-BRCT domain. J
Mol Biol. 2013 Sep 9;425(17):3091-105. doi: 10.1016/j.jmb.2013.05.029. Epub 2013
Jun 7. PubMed PMID: 23747975.
14: Korzhnev DM. Loss of structure-gain of function. J Mol Biol. 2013 Jan
9;425(1):17-8. doi: 10.1016/j.jmb.2012.10.012. Epub 2012 Oct 22. PubMed PMID:
23084974.
15: Markley JL, Akutsu H, Asakura T, Baldus M, Boelens R, Bonvin A, Kaptein R,
Bax A, Bezsonova I, Gryk MR, Hoch JC, Korzhnev DM, Maciejewski MW, Case D, Chazin
WJ, Cross TA, Dames S, Kessler H, Lange O, Madl T, Reif B, Sattler M, Eliezer D,
Fersht A, Forman-Kay J, Kay LE, Fraser J, Gross J, Kortemme T, Sali A, Fujiwara
T, Gardner K, Luo X, Rizo-Rey J, Rosen M, Gil RR, Ho C, Rule G, Gronenborn AM,
Ishima R, Klein-Seetharaman J, Tang P, van der Wel P, Xu Y, Grzesiek S, Hiller S,
Seelig J, Laue ED, Mott H, Nietlispach D, Barsukov I, Lian LY, Middleton D,
Blumenschein T, Moore G, Campbell I, Schnell J, Vakonakis IJ, Watts A, Conte MR,
Mason J, Pfuhl M, Sanderson MR, Craven J, Williamson M, Dominguez C, Roberts G,
Günther U, Overduin M, Werner J, Williamson P, Blindauer C, Crump M, Driscoll P,
Frenkiel T, Golovanov A, Matthews S, Parkinson J, Uhrin D, Williams M, Neuhaus D,
Oschkinat H, Ramos A, Shaw DE, Steinbeck C, Vendruscolo M, Vuister GW, Walters
KJ, Weinstein H, Wüthrich K, Yokoyama S. In support of the BMRB. Nat Struct Mol
Biol. 2012 Sep;19(9):854-60. doi: 10.1038/nsmb.2371. PubMed PMID: 22955930.
16: Pustovalova Y, Bezsonova I, Korzhnev DM. The C-terminal domain of human Rev1
contains independent binding sites for DNA polymerase η and Rev7 subunit of
polymerase ζ. FEBS Lett. 2012 Sep 21;586(19):3051-6. doi:
10.1016/j.febslet.2012.07.021. Epub 2012 Jul 22. PubMed PMID: 22828282; PubMed
Central PMCID: PMC3572780.
17: Pozhidaeva A, Pustovalova Y, D'Souza S, Bezsonova I, Walker GC, Korzhnev DM.
NMR structure and dynamics of the C-terminal domain from human Rev1 and its
complex with Rev1 interacting region of DNA polymerase η. Biochemistry. 2012 Jul
10;51(27):5506-20. Epub 2012 Jun 28. PubMed PMID: 22691049; PubMed Central PMCID:
PMC3732116.
18: Korzhnev DM, Religa TL, Kay LE. Transiently populated intermediate functions
as a branching point of the FF domain folding pathway. Proc Natl Acad Sci U S A.
2012 Oct 30;109(44):17777-82. doi: 10.1073/pnas.1201799109. Epub 2012 May 30.
PubMed PMID: 22647611; PubMed Central PMCID: PMC3497754.
19: Barette J, Velyvis A, Religa TL, Korzhnev DM, Kay LE. Cross-validation of the
structure of a transiently formed and low populated FF domain folding
intermediate determined by relaxation dispersion NMR and CS-Rosetta. J Phys Chem
B. 2012 Jun 14;116(23):6637-44. doi: 10.1021/jp209974f. Epub 2011 Dec 23. PubMed
PMID: 22148426.
20: Korzhnev DM, Vernon RM, Religa TL, Hansen AL, Baker D, Fersht AR, Kay LE.
Nonnative interactions in the FF domain folding pathway from an atomic resolution
structure of a sparsely populated intermediate: an NMR relaxation dispersion
study. J Am Chem Soc. 2011 Jul 20;133(28):10974-82. doi: 10.1021/ja203686t. Epub
2011 Jun 28. PubMed PMID: 21639149; PubMed Central PMCID: PMC3705915.
21: Korzhnev DM, Religa TL, Banachewicz W, Fersht AR, Kay LE. A transient and
low-populated protein-folding intermediate at atomic resolution. Science. 2010
Sep 10;329(5997):1312-6. doi: 10.1126/science.1191723. PubMed PMID: 20829478.
22: Fatemi N, Korzhnev DM, Velyvis A, Sarkar B, Forman-Kay JD. NMR
characterization of copper-binding domains 4-6 of ATP7B . Biochemistry. 2010 Oct
5;49(39):8468-77. doi: 10.1021/bi1008535. Epub 2010 Sep 10. PubMed PMID:
20799727.
23: Bouvignies G, Korzhnev DM, Neudecker P, Hansen DF, Cordes MH, Kay LE. A
simple method for measuring signs of (1)H (N) chemical shift differences between
ground and excited protein states. J Biomol NMR. 2010 Jun;47(2):135-41. doi:
10.1007/s10858-010-9418-8. Epub 2010 Apr 29. PubMed PMID: 20428928; PubMed
Central PMCID: PMC3034452.
24: Auer R, Hansen DF, Neudecker P, Korzhnev DM, Muhandiram DR, Konrat R, Kay LE.
Measurement of signs of chemical shift differences between ground and excited
protein states: a comparison between H(S/M)QC and R1rho methods. J Biomol NMR.
2010 Mar;46(3):205-16. doi: 10.1007/s10858-009-9394-z. Epub 2009 Dec 22. PubMed
PMID: 20033258.
25: van Ingen H, Korzhnev DM, Kay LE. An analysis of the effects of 1HN-(1)HN
dipolar couplings on the measurement of amide bond vector orientations in
invisible protein states by relaxation dispersion NMR. J Phys Chem B. 2009 Jul
23;113(29):9968-77. doi: 10.1021/jp902793y. PubMed PMID: 19569643.
…
26: Korzhnev DM, Bezsonova I, Lee S, Chalikian TV, Kay LE. Alternate binding
modes for a ubiquitin-SH3 domain interaction studied by NMR spectroscopy. J Mol
Biol. 2009 Feb 20;386(2):391-405. doi: 10.1016/j.jmb.2008.11.055. Epub 2008 Dec
6. PubMed PMID: 19111555.
27: Korzhnev DM, Kay LE. Probing invisible, low-populated States of protein
molecules by relaxation dispersion NMR spectroscopy: an application to protein
folding. Acc Chem Res. 2008 Mar;41(3):442-51. doi: 10.1021/ar700189y. Epub 2008
Feb 15. Review. PubMed PMID: 18275162.
28: Murphy JM, Korzhnev DM, Ceccarelli DF, Briant DJ, Zarrine-Afsar A, Sicheri F,
Kay LE, Pawson T. Conformational instability of the MARK3 UBA domain compromises
ubiquitin recognition and promotes interaction with the adjacent kinase domain.
Proc Natl Acad Sci U S A. 2007 Sep 4;104(36):14336-41. Epub 2007 Aug 28. PubMed
PMID: 17726107; PubMed Central PMCID: PMC1964837.
29: Korzhnev DM, Religa TL, Lundström P, Fersht AR, Kay LE. The folding pathway
of an FF domain: characterization of an on-pathway intermediate state under
folding conditions by (15)N, (13)C(alpha) and (13)C-methyl relaxation dispersion
and (1)H/(2)H-exchange NMR spectroscopy. J Mol Biol. 2007 Sep 14;372(2):497-512.
Epub 2007 Jun 9. PubMed PMID: 17689561.
30: Zhuravleva A, Korzhnev DM, Nolde SB, Kay LE, Arseniev AS, Billeter M, Orekhov
VY. Propagation of dynamic changes in barnase upon binding of barstar: an NMR and
computational study. J Mol Biol. 2007 Apr 6;367(4):1079-92. Epub 2007 Jan 24.
PubMed PMID: 17306298.
31: Korzhnev DM, Neudecker P, Zarrine-Afsar A, Davidson AR, Kay LE. Abp1p and Fyn
SH3 domains fold through similar low-populated intermediate states. Biochemistry.
2006 Aug 29;45(34):10175-83. PubMed PMID: 16922492.
32: Korzhnev DM, Bezsonova I, Evanics F, Taulier N, Zhou Z, Bai Y, Chalikian TV,
Prosser RS, Kay LE. Probing the transition state ensemble of a protein folding
reaction by pressure-dependent NMR relaxation dispersion. J Am Chem Soc. 2006 Apr
19;128(15):5262-9. PubMed PMID: 16608362.
33: Bezsonova I, Korzhnev DM, Prosser RS, Forman-Kay JD, Kay LE. Hydration and
packing along the folding pathway of SH3 domains by pressure-dependent NMR.
Biochemistry. 2006 Apr 18;45(15):4711-9. PubMed PMID: 16605239.
34: Neudecker P, Korzhnev DM, Kay LE. Assessment of the effects of increased
relaxation dispersion data on the extraction of 3-site exchange parameters
characterizing the unfolding of an SH3 domain. J Biomol NMR. 2006
Mar;34(3):129-35. PubMed PMID: 16604422.
35: Mittermaier A, Korzhnev DM, Kay LE. Side-chain interactions in the folding
pathway of a Fyn SH3 domain mutant studied by relaxation dispersion NMR
spectroscopy. Biochemistry. 2005 Nov 29;44(47):15430-6. PubMed PMID: 16300390.
36: Eisenmesser EZ, Millet O, Labeikovsky W, Korzhnev DM, Wolf-Watz M, Bosco DA,
Skalicky JJ, Kay LE, Kern D. Intrinsic dynamics of an enzyme underlies catalysis.
Nature. 2005 Nov 3;438(7064):117-21. PubMed PMID: 16267559.
37: Korzhnev DM, Neudecker P, Mittermaier A, Orekhov VY, Kay LE. Multiple-site
exchange in proteins studied with a suite of six NMR relaxation dispersion
experiments: an application to the folding of a Fyn SH3 domain mutant. J Am Chem
Soc. 2005 Nov 9;127(44):15602-11. PubMed PMID: 16262426.
38: Korzhnev DM, Mittermaier AK, Kay LE. Cross-correlated spin relaxation effects
in methyl 1H CPMG-based relaxation dispersion experiments: complications and a
simple solution. J Biomol NMR. 2005 Apr;31(4):337-42. PubMed PMID: 15929000.
39: Korzhnev DM, Orekhov VY, Kay LE. Off-resonance R(1rho) NMR studies of
exchange dynamics in proteins with low spin-lock fields: an application to a Fyn
SH3 domain. J Am Chem Soc. 2005 Jan 19;127(2):713-21. PubMed PMID: 15643897.
40: Zhuravleva AV, Korzhnev DM, Kupce E, Arseniev AS, Billeter M, Orekhov VY.
Gated electron transfers and electron pathways in azurin: a NMR dynamic study at
multiple fields and temperatures. J Mol Biol. 2004 Oct 1;342(5):1599-611. PubMed
PMID: 15364584.
41: Korzhnev DM, Salvatella X, Vendruscolo M, Di Nardo AA, Davidson AR, Dobson
CM, Kay LE. Low-populated folding intermediates of Fyn SH3 characterized by
relaxation dispersion NMR. Nature. 2004 Jul 29;430(6999):586-90. PubMed PMID:
15282609.
42: Korzhnev DM, Kloiber K, Kay LE. Multiple-quantum relaxation dispersion NMR
spectroscopy probing millisecond time-scale dynamics in proteins: theory and
application. J Am Chem Soc. 2004 Jun 16;126(23):7320-9. PubMed PMID: 15186169.
43: Di Nardo AA, Korzhnev DM, Stogios PJ, Zarrine-Afsar A, Kay LE, Davidson AR.
Dramatic acceleration of protein folding by stabilization of a nonnative backbone
conformation. Proc Natl Acad Sci U S A. 2004 May 25;101(21):7954-9. Epub 2004 May
17. PubMed PMID: 15148398; PubMed Central PMCID: PMC419538.
44: Korzhnev DM, Kloiber K, Kanelis V, Tugarinov V, Kay LE. Probing slow dynamics
in high molecular weight proteins by methyl-TROSY NMR spectroscopy: application
to a 723-residue enzyme. J Am Chem Soc. 2004 Mar 31;126(12):3964-73. PubMed PMID:
15038751.
45: Bocharov EV, Sobol AG, Pavlov KV, Korzhnev DM, Jaravine VA, Gudkov AT,
Arseniev AS. From structure and dynamics of protein L7/L12 to molecular switching
in ribosome. J Biol Chem. 2004 Apr 23;279(17):17697-706. Epub 2004 Feb 11. PubMed
PMID: 14960595.
46: Orekhov VY, Korzhnev DM, Kay LE. Double- and zero-quantum NMR relaxation
dispersion experiments sampling millisecond time scale dynamics in proteins. J Am
Chem Soc. 2004 Feb 18;126(6):1886-91. PubMed PMID: 14871121.
47: Korzhnev DM, Orekhov VY, Dahlquist FW, Kay LE. Off-resonance R1rho relaxation
outside of the fast exchange limit: an experimental study of a cavity mutant of
T4 lysozyme. J Biomol NMR. 2003 May;26(1):39-48. PubMed PMID: 12766401.
48: Korzhnev DM, Karlsson BG, Orekhov VY, Billeter M. NMR detection of multiple
transitions to low-populated states in azurin. Protein Sci. 2003 Jan;12(1):56-65.
PubMed PMID: 12493828; PubMed Central PMCID: PMC2312403.
49: Bocharov EV, Korzhnev DM, Blommers MJ, Arvinte T, Orekhov VY, Billeter M,
Arseniev AS. Dynamics-modulated biological activity of transforming growth factor
beta3. J Biol Chem. 2002 Nov 29;277(48):46273-9. Epub 2002 Sep 6. PubMed PMID:
12221089.
50: Korzhnev DM, Skrynnikov NR, Millet O, Torchia DA, Kay LE. An NMR experiment
for the accurate measurement of heteronuclear spin-lock relaxation rates. J Am
Chem Soc. 2002 Sep 11;124(36):10743-53. PubMed PMID: 12207529.
51: Korzhnev DM, Bocharov EV, Zhuravlyova AV, Orekhov VY, Ovchinnikova TV,
Billeter M, Arseniev AS. Backbone dynamics of the channel-forming antibiotic
zervamicin IIB studied by 15N NMR relaxation. FEBS Lett. 2001 Apr
20;495(1-2):52-5. PubMed PMID: 11322946.
52: Korzhnev DM, Tischenko EV, Arseniev AS. Off-resonance effects in 15N T2 CPMG
measurements. J Biomol NMR. 2000 Jul;17(3):231-7. PubMed PMID: 10959630.
53: Orekhov VY, Korzhnev DM, Pervushin KV, Hoffmann E, Arseniev AS. Sampling of
protein dynamics in nanosecond time scale by 15N NMR relaxation and
self-diffusion measurements. J Biomol Struct Dyn. 1999 Aug;17(1):157-74. PubMed
PMID: 10496429.
54: Korzhnev DM, Orekhov VY, Arseniev AS. Model-free approach beyond the borders
of its applicability. J Magn Reson. 1997 Aug;127(2):184-91. PubMed PMID: 9281482.
55: Orekhov VY, Pervushin KV, Korzhnev DM, Arseniev AS. Backbone dynamics of
(1-71)- and (1-36)bacterioopsin studied by two-dimensional (1)H- (15)N NMR
spectroscopy. J Biomol NMR. 1995 Sep;6(2):113-22. doi: 10.1007/BF00211774. PubMed
PMID: 22910799.
56: Pervushin KV, Orekhov VY, Korzhnev DM, Arseniev AS. Manifestation of
intramolecular motions on pico- and nanosecond time scales in (1)H- (15)N NMR
relaxation: Analysis of dynamic models of one- and two-helical subunits of
bacterioopsin. J Biomol NMR. 1995 Jun;5(4):383-96. doi: 10.1007/BF00182282.
PubMed PMID: 22911558.