The research in my laboratory has three major directions: to elucidate kinetic mechanisms of enzymes involved in DNA/RNA replication, repair, and lesion bypass; to understand Hepatitis C (HCV) replication and regulation of innate immunity; to develop antiviral and anti-cancer molecules based on rational drug design. 

In the Suo lab, pre-steady state kinetic methods are employed using rapid chemical quench-flow and stopped-flow. These methods allow us to quench reactions on the millisecond time scale and to extract more kinetic information than the traditional steady-state kinetic methods. We also use protein engineering methods including site-directed mutagenesis and domain-swapping to study structure-function relationships of DNA polymerases. Recently, we have initiated projects to investigate the dynamics of a DNA polymerase using NMR and single-molecule techniques. Moreover, we are using femtosecond-resolved fluorescence up-conversion techniques to study enzyme-substrate interactions in collaboration with the research group of Dr. Dongping Zhong in the Department of Physics at OSU. X-ray crystallography will be used to examine interesting enzyme-substrate complexes. These multi-disciplinary approaches will allow us to develop new methods and to advance enzymology into unprecedented territory. Our goals are to understand the elementary steps of conformational changes and chemical reactions occurring at the active site of enzymes. Then, these mechanisms will be used for rational drug design. The designed enzyme inhibitors will be synthesized and tested in vitro and in vivo. Currently, we are investigating several systems described below.

(1) Pre-Steady State Kinetic Studies of DNA Lesion Bypass Polymerases

DNA lesions often block DNA replication, so cells possess specific, often error-prone, DNA polymerases to bypass such lesions and to promote replication of damaged DNA. More than 220 DNA lesion bypass polymerases have been discovered. These polymerases, which share sequence similarity and catalyze DNA polymerization with low fidelity and poor processivity, are classified into a new family: the Y-family. Human polymerases eta (h), iota (i), kappa (k) and Rev1 are examples of DNA lesion bypass enzymes. Pol h, encoded by hRAD30A, bypasses cis-syn thymine-thymine dimers efficiently and accurately. Mutations in hRAD30A inactivate Pol h and lead to UV-induced mutagenesis and skin cancer. The Suo laboratory is using pre-steady state kinetic methods to decipher the detailed mechanisms of correct and incorrect nucleotide incorporations opposite undamaged or damaged DNA templates by Dpo4, a thermostable polymerase from Sulfolobus solfataricus strain P2, Pol h, Pol i, Pol k, and Rev1. We have developed a novel assay, short oligonucleotide sequencing assay (SOSA), to determine the DNA sequence of lesion bypass products synthesized by Y-family enzymes. We are collaborating with the group of Dr. Hong Ling at the University of Western Ontario, Canada, to crystallize the binary and ternary complexes of Dpo4. We are also collaborating with the group of Dr. Dongping Zhong to study the dynamic interactions among Dpo4, DNA, and an incoming nucleotide using the femtosecond-resolved fluorescence up-conversion techniques. Our collaborative studies will establish a general kinetic, thermodynamic, and structural mechanism for DNA translesion synthesis. More importantly, a better knowledge of the Y-family polymerases based on our results will facilitate the understanding of cancer formation and the development of anticancer drugs.

(2) Kinetic and Protein-Protein Interaction Studies of Human DNA Polymerases lambda, mu and TdT

The DNA in every cell of the human body is spontaneously damaged more than 10,000 times every day. DNA repair plays a major role in maintaining the integrity of genomic DNA in cells. Human DNA polymerase lambda (l) shares sequence similarity with the well-known DNA repair polymerase beta (b) and is thereby believed to catalyze base excision repair. Human DNA polymerase mu (µ) shares sequence and functions similar with the well-known human deoxynucleotidyl transferase (TdT) which participates in antibody generation. My group has purified three X-family polymerases and is employing pre-steady state kinetic methods to characterize their kinetic mechanisms. In addition, Pol l and Pol µ have an N-terminal BRCT domain which interacts with cell-cycle checking proteins, such as the tumor suppressor p53. We are trying to identify these interacting proteins by employing immuno-precipitation assay and mass spectroscopy analysis. Moreover, we are trying to crystallize both Pol l and Pol µ in the presence of DNA and dNTP substrates.

(3)  Mechanistic Studies of Vaccinia Virus DNA Polymerases and Design/Synthesis of Novel Nucleoside Analog Inhibitors

Concerns about the possible release of smallpox by bioterrorists have led to intensive hunt to find an effective molecule to inhibit viral infection which does not exist yet. Since smallpox virus (variola virus) and the smallpox vaccine (vaccinia virus) are highly homologous, the latter has been used as a very good surrogate model. For example, vaccinia virus DNA polymerase is about 99% identical to its counterpart in smallpox virus. In my laboratory, we are using pre-steady state kinetic methods to investigate the elementary steps of nucleotide incorporation catalyzed by vaccinia virus DNA polymerase. In addition, we are testing more than 140 nucleotide analogs in order to find potent inhibitors which may be effective as anti-smallpox agents.

(4)  Design and Synthesis of Novel Nucleoside Analog Inhibitors

Hepatitis C has infected about 2-3% of human population. Viral genome replication is crucial for viral life cycles and has been studied intensively. NS5B, the RNA-dependant RNA polymerase, which is at the center of viral replication, is one of major antiviral drug targets. Although there are extensive biochemical and steady-state kinetic studies on this polymerase, the elementary steps of nucleotide incorporation catalyzed by NS5B are still undefined. Using pre-steady state kinetic methods, we are studying the kinetic mechanism, processivity, fidelity, drug susceptibility, and drug resistance. The knowledge gained from these studies has severed as the basis for our rational design of nucleoside inhibitors. Currently, we are testing more than 140 nucleoside analogs which we have synthesized or obtained through collaboration in our cell-based assays.

(5) Developing Anti-HCV Peptide-Based Inhibitors

The non-structural proteins NS3, NS4A, NS4B, NS5A, and NS5B of HCV are processed from viral polyprotein precursor C-E1-E2-p7-NS2-NS3-NS4A-NS4B-NS5A-NS5B by viral protease complex NS3/NS4A. NS3 has an N-terminal protease domain and a C-terminal helicase domain. The crystal structure of the NS3 protease domain shows that the N-terminus 28 residues are unfolded. In the complex with NS4A, the NS3 N-terminus folds into a beta sheet and an alpha helix, and the active site residues are slightly rearranged to form a catalytically favorable conformation. The NS3 protease is 995-fold more active in the presence than in the absence of NS4A. We are using the Stopped-Flow technology to study these conformational changes in NS3 after NS4A binding. We are also searching for tighter binding peptides to inhibit NS4A binding to NS3. The peptide inhibitors are then tested in the liver cell-line Huh 7-based HCV replicon assay. The inhibitory mechanism of the best peptide inhibitors will be studied further use confocal and multiphoton imaging and microscopy.

(6)  Effects of HCV Protease NS3/4A on Human Kinases Involved in Immune Response

Virus infection signals antiviral response through transcription factors, nuclear factor k-B (NFkB) and interferon regulatory factors (IRFs). Current treatment includes interferon-a (IFN-a) based therapy that amplifies host antiviral response. In contrast, HCV has evolved unknown mechanisms to disrupt the host response to IFN-a. To examine the effect of HCV protease NS3/4A on these pathways, we are collaborating with Dr. T. Maniatis at Harvard University to elucidate these novel pathways.

Ma, D., Fowler, J.D., and Suo Z.*(2011) Backbone Assignment of the Little Finger Domain of a Y-Family DNA Polymerase, Biomolecular NMR Assignments. In press.

Kirouac, K. N., Suo, Z., and H. Ling* (2011) Structural mechanism of ribonucleotide discrimination by a Y family DNA polymerase, J. Mol. Biol. In press.

Brown, J.A. and Suo, Z.* (2011) Unlocking the Sugar 'Steric Gate' of DNA Polymerases, Biochemistry, invited review 50, 1135-1142.

Brown, J.A., Pack, L.R., Fowler, J.D. and Suo Z.* (2011) Pre-Steady State Kinetic Analysis of the Incorporation of Anti-HIV Nucleotide Analogs Catalyzed by Human X- and Y-family DNA Polymerases, Antimicrob. Agents Chemother. 55, 276-283.

Brown, J.A., Pack, L.R., Sanman, L.E. and Suo Z.* (2011) Efficiency and Fidelity of Human DNA Polymerases Lambda and Beta during Gap-Filling DNA Synthesis, DNA Repair 10, 24-33.

Sherrer S.M., Fiala, K.A., Fowler J.D., Newmister, S.A., Pryor, J.M. and Suo Z.* (2011) Quantitative Analysis of the Efficiency and Mutagenic Spectra of Abasic Lesion Bypass Catalyzed by Human Y-family DNA Polymerases, Nucleic Acids Research 39, 609–622.

Sherrer S.M., Beyer, D.C., Xia, C.X., Fowler J.D., and Suo Z.* (2010) Kinetic Basis of Sugar Selection by a Y-family DNA Polymerase from Sulfolobus solfataricus P2, Biochemistry 49, 10179-10186.

Brown, J.A., Pack, L.R., Sherrer, S.M., Kshetry, A.K., Newmister, S.A., Fowler, J.D., Taylor, J.-S. and Suo Z.* (2010) Identification of Critical Residues for the Tight Binding of Both Correct and Incorrect Nucleotides to Human DNA Polymerase Lambda, J. Mol. Biol. 403, 505-515.

Ma, D., Fowler, J.D., Yuan, C. and Suo Z.* (2010) Backbone Assignment of the Catalytic Core of a Y-family DNA Polymerase, Biomolecular NMR Assignments 4, 207-209 DOI: 10.1007/s12104-010-9244-7.

Brown, J.A., Fowler, J.D. and Suo Z.* (2010) Kinetic Basis of Nucleotide Selection Employed by a Protein Template-Dependent DNA Polymerase, Biochemistry 49, 5504-5510.

Wong, J.H., Brown, J.A., Suo Z., Blum, P., Nohmi, T. and Ling, H.* (2010) Structural Insight into Dynamic Bypass of the Major Cisplatin-DNA Adduct by Y-family Polymerase Dpo4, EMBO J. 29, 2059-2069.

Brown, J.A., Zhang, L., Sherrer, S.M., Taylor, J.-S., Burgers, P.M.J. and Suo Z.* (2010) Pre-Steady State Kinetic Analysis of Truncated and Full-Length Saccharomyces cerevisiae DNA Polymerase Eta, J. Nucleic Acids 2010, 11 pages.

Brown, J.A., Fiala, K.A., Fowler, J.D., Sherrer, S.M., Newmister, S.A., Duym, W.W. and Suo Z.* (2010) A Novel Mechanism of Sugar Selection Utilized by a Human X-family DNA Polymerase, J. Mol. Biol. 395, 282-290.

Xu, C., Maxwell, B.A., Brown, J.A., Zhang, L. and Suo Z.* (2009) Global Conformational Dynamics of a Y-family DNA Polymerase during Catalysis, PLoS Biology 7(10): e1000225.

Brown, J.A. and Suo Z.* (2009) Elucidating the Kinetic Mechanism of DNA Polymerization Catalyzed by Sulfolobus solfataricus P2 DNA Polymerase B1, Biochemistry 48, 7502-7511.

Zhang, L., Brown, J.A., Newmister, S.A. and Suo Z.* (2009) Polymerization Fidelity of a Replicative DNA Polymerase from the Hyperthermophilic Archaeon Sulfolobus solfataricus P2, Biochemistry 48, 7492-7501.

Fowler J.D., Brown, J.A., Kvaratskhelia, M. and Suo Z.* (2009) Probing Conformational Changes of Human DNA Polymerase Lambda using Mass Spectrometry-Based Protein Footprinting, J. Mol. Biol. 390, 368-379.

Sherrer S.M., Brown J.A., Pack L.R., Jasti V.P., Fowler J.D., Basu A.K., and Suo Z.* (2009) Mechanistic Studies of the Bypass of a Bulky Single-Base Lesion Catalyzed by a Y-family DNA Polymerase, J. Biol. Chem. 284, 6379-8863.

DeCarlo, L., Prakasha Gowda, A.S., Suo, Z. and Spratt, T.E.* (2008) Formation of Purine-Purine Mispairs by Sulfolobus solfataricus DNA Polymerase IV, Biochemistry 47, 8157-8164.

Brown, J.A., Newmister, S.A., Fiala, K.A. and Suo, Z.* (2008) Mechanism of Double-Base Lesion Bypass Catalyzed by a Y-family DNA Polymerase, Nucleic Acids Research 36, 3867-3878.

Wong, J.H., Fiala, K.A., Suo, Z. and Ling, H.* (2008) Snapshots of a Y-family DNA Polymerase in Replication: Substrate-Induced Conformational Transitions and Implications for Fidelity of Dpo4, J. Mol. Biol. 379, 317-330.

Fowler, J.D., Brown, J.A., Johnson, K.A. and Suo, Z.* (2008) Kinetic Investigation of the Inhibitory Effect of Gemcitabine on DNA Polymerization Catalyzed by Human Mitochondrial DNA Polymerase. J. Biol. Chem. 283, 15339-15348.

Fiala, K.A., Sherrer, S.M., Brown, J.A. and Suo, Z.* (2008) Mechanistic Consequences of Temperature on DNA Polymerization Catalyzed by a Y-family DNA Polymerase, Nucleic Acids Research 36, 1990-2001.

Brown JA, Duym WW, Fowler JD, Suo, Z.. (2007) "Single-turnover Kinetic Analysis of the Mutagenic Potential of 8-Oxo-7,8-dihydro-2'-deoxyguanosine during Gap-filling Synthesis Catalyzed by Human DNA Polymerases lambda and beta." J Mol Biol. 367, 1258-1269

Suo, Z., Abdullah MA. (2007) "Unique Composite Active Site of the Hepatitis C Virus NS2-3 Protease: a New Opportunity for Antiviral Drug Design." ChemMedChem. 2(3), 283-284.

Fiala KA, Suo, Z. (2007) "Sloppy bypass of an abasic lesion catalyzed by a Y-family DNA polymerase." J Biol Chem. 282, 8199-8206

Fiala KA, Hypes CD, Suo, Z. (2007) "Mechanism of abasic lesion bypass catalyzed by a Y-family DNA polymerase." J Biol Chem. 282, 8188-8198

Fiala KA, Brown JA, Ling H, Kshetry AK, Zhang J, Taylor JS, Yang W, Suo, Z.. (2007) "Mechanism of template-independent nucleotide incorporation catalyzed by a template-dependent DNA polymerase." J Mol Biol. 365(3), 590-602.

Duym WW, Fiala KA, Bhatt N, Suo, Z.. (2006) "Kinetic effect of a downstream strand and its 5'-terminal moieties on single nucleotide gap-filling synthesis catalyzed by human DNA polymerase lambda." J Biol Chem. 281(47), 35649-55.

Fowler JD, Suo, Z.. (2006) "Biochemical, structural, and physiological characterization of terminal deoxynucleotidyl transferase." Chem Rev. 106(6), 2092-110.

Fiala KA, Duym WW, Zhang J, Suo, Z.. (2006) "Up-regulation of the fidelity of human DNA polymerase lambda by its non-enzymatic proline-rich domain." J Biol Chem. 281(28), 19038- 44.

Suo Z.  (2005) "Thioesterase portability and peptidyl carrier protein swapping in yersiniabactin synthetase from Yersinia pestis.", Biochemistry 44(12), 4926-38.

Roettger MP, Fiala KA, Sompalli S, Dong Y, Suo Z.  (2004) "Pre-steady-state kinetic studies of the fidelity of human DNA polymerase mu", Biochemistry 43(43), 13827-38.

Fiala KA, Abdel-Gawad W, Suo Z.  (2004) "Pre-steady-state kinetic studies of the fidelity and mechanism of polymerization catalyzed by truncated human DNA polymerase lambda.", Biochemistry 43(21), 6751-62.

Fiala, K. A & Suo Z.* (2004) Pre-Steady State Kinetic Studies of the Fidelity of Sulfolobus solfataricus P2 DNA Polymerase IV.Biochemistry 43, 2106-2115

Fiala, K. A & Suo Z.* (2004) Mechanism of DNA Polymerization Catalyzed by Sulfolobus solfataricus P2 DNA Polymerase IV. Biochemistry 43, 2116-2125

Zhang G. & Suo Z.* (2004) A Mild and Convenient Synthetic Method for Arylhydrazones of Methyl Benzoate. Synthetic Communications 34(4), 673-678.

Fiala, K. A, Abdel-Gawad, W. & Suo Z.* (2004) Pre-Steady-State Kinetic Studies of the Fidelity and Mechanism of Polymerization Catalyzed by Truncated Human DNA Polymerase Lambda.Biochemistry, accepted and in press.

Allison, A. J., Ray, A., Suo Z.., Colacino, J. M., Andeson, K. S., Johnson, K.A. (2001) “Toxicity of Antiviral Nucleoside Analogs and the Human Mitochondrial DNA Polymerase", J. Biol. Chem. 276, 40847-40857.

Suo Z.. & Walsh, C. T. (2001) “Thioesterase Portability and Peptidyl Carrier Protein Swapping in Yersiniabactin Synthetase from Yersinia pestis”, Biochemistry, submitted.

Suo Z.., Tseng, C., & Walsh, C. T. (2001) “Purification, Priming, and Catalytic Acylation of Carrier Protein Domains in the Polyketide Synthase and Nonribosomal Peptidyl Synthetase Modules of the HMWP1 Subunit of Yersiniabactin Synthetase”, Proc. Natl. Acad. Sci. U.S.A. 98, 99-104.