Istvan Molnar
Associate Professor
Title:
Associate ProfessorArea of Expertise:
Biosynthetic engineering, microbial genetics, combinatorial biosynthesis, genomics, biocatalysis, industrial biotechnology, drug discovery, natural products, antibiotics, anticancer agents, biofuels
Dr Molnar is an Associate Professor at the Natural Products Center, and an adjunct at the Division of Plant Pathology and Microbiology of the School of Plant Sciences. He is also a Program Faculty at the Microbiology Graduate Program, the Arid Lands Resource Sciences GIDP, the Arid Lands Sustainable Bio-Energy Institute, and the Bio5 Institute for Collaborative Bioresearch.
My main research interest is the biosynthetic engineering of microorganisms (bacteria, fungi and algae) for pharmaceutical, agricultural, chemical industrial and bioenergy applications. My research applies modern microbiology, microbial genetics and biochemistry, genomics, metabolic engineering, microbial systems biology, synthetic biology, and a dash of chemistry to the understanding of the biosynthetic processes of these organisms. My main focus is on secondary metabolism, the biosynthesis of small molecule natural products that are used as antibiotics, anticancer agents and other drugs in human medicine, veterinary practice, and agriculture. I am also interested in applying the same methodologies to devise sustainable processes for the production of carbon-neutral, renewable energy in the form of biofuels.
Microorganisms have practiced wonderfully elegant and incredibly complex (bio)chemistry to produce structurally complex secondary metabolites for hundreds of millions of years. These metabolites represent privileged structures selected by evolution to interact with many, possibly all, protein folds in nature. They show not only exquisite specificity to their targets, but are generally stable in biological environments, and often able to penetrate cells to interact with intracellular targets. Secondary metabolites are agents of competition and chemical warfare in microbial communities, but they also represent virulence factors in pathogenic contexts, and carry biological information amongst cells to coordinate their behavior. Utilizing these complex (and to me, beautiful) molecules for human purposes represents a major accomplishment of the pharmaceutical industry and the academic research community. Natural products have contributed well over 10 years to our expected lifespan as life saving drugs, and help us feed our growing numbers by serving as veterinary and agricultural agents.
Continued exploitation of natural products requires us to be able to modify these small molecules to even better suit our needs. While synthetic and medicinal chemistry provides true-and-tested methods to produce natural product analogues and derivatives, utilizing the microorganisms themselves to synthesize these “humanized” molecules is still an attractive proposition. Biosynthesis can provide processes that can be described as “green chemistry”: complex reaction sequences without toxic chemicals, conducted under mild conditions. Biosynthesis offers scalable and sustainable production processes using cheap feedstocks, and promises substantial savings in production costs.
Understanding how microorganisms produce natural products provides a blueprint that might be edited or re-drawn to affect the biosynthesis of modified “unnatural” products. Deciphering the master plan of cellular metabolic processes, as revealed by different “omics” methods, will also allow us to channel a larger fraction of the resources of the bioengineered cells towards synthesizing larger amounts of economically important molecules like natural products or biofuels.
After doing research in both academic institutions and the international biotech industry, I joined the UA at the end of 2004. Here, I evolved from studying my beloved bacteria to also experimenting with filamentous fungi, and became interested in engineering algae.
My group cloned and analyzed the biosynthetic gene clusters for two fungal nonribosomal depsipeptides, beauvericin and bassianolide. Beauvericin has potent toxicity to cancer cells, and also inhibits cell motility, potentially reducing tumor growth and metastasis. We worked out a heterologous expression system for beauvericin in E. coli, a first in producing such fungal metabolites in a domesticated bacterial host. In collaboration with Prof. Gunatilaka’s group at the Natural Product Center, we produced a variety of beauvericin analogues by combinatorial biosynthesis, and determined the effects of the structural changes on the biological activity of these “unnatural” natural products. In collaboration with Prof. Stock’s group at the Department of Entomology, we have established that both beauvericin and bassianolide are used by the producer fungus, Beauveria bassiana, as virulence factors during fungal infection of insects.
In another project, we cloned and analyzed the biosynthetic gene cluster for radicicol, a fungal heat shock protein inhibitor. Radicicol displays potent anticancer activities, but its analogues also increase the heat stress tolerance of plants. We have shown that the radicicol scaffold is produced by two collaborating fungal polyketide synthase (PKS) enzymes, a rather unusual arrangement in fungal polyketide biosynthesis. We have discovered that radicicol biosynthesis also involves a FAD-dependent chlorinase enzyme: such substrate- and regiospecific halogenases have not previously been identified in fungi. We have also produced radicicol analogues by targeted gene inactivation in the producer fungus, Chaetomium chiversii.
To remain true to my prokaryotic roots, we have investigated a library of environmental bacteria for natural products with potential anticancer activities. These strains were collected from the Sonoran desert by Prof. Gunatilaka of the Natural Products Center and his collaborators at the Plant Pathology and Microbiology.
I am also taking part in a new collaborative initiative at the UA that aims to utilize algae for the production of metabolites that might be used as biofuels. My interest is the characterization and engineering of the relevant biosynthetic pathways that are currently not well understood. This UA collaboration is a funding member of the National Alliance for Advanced Biofuels and Bioproducts (NAABB) that has recently received a major grant from the Department of Energy. NAABB intends to develop renewable, carbon-neutral advanced biodiesel and biojetfuel products that can be mass-produced for the transporation needs of the nation.
Publications:
1. Coleman, J.J., Rounsley, S.D., Rodriguez-Carres, M., Kuo, A., Wasmann, C.C., Grimwood, J., Schmutz, J., Taga, M., White, G.J., Zhou, S., Schwartz, D.C., Freitag, M., Ma, L.J., Danchin, E.G., Henrissat, B., Coutinho, P.M., Nelson, D.R., Straney, D., Napoli, C.A., Barker, B.M., Gribskov, M., Rep, M., Kroken, S., Molnar, I., Rensing, C., Kennell, J.C., Zamora, J., Farman, M.L., Selker, E.U., Salamov, A., Shapiro, H., Pangilinan, J., Lindquist, E., Lamers, C., Grigoriev, I.V., Geiser, D.M., Covert, S.F., Temporini, E., VanEtten, H.D.: The genome of Nectria haematococca: contribution of supernumerary chromosomes to gene expansion. PLOS Genet. 2009;5: e1000618.
2. Xu, Y., Orozco, R., Wijeratne, E.M.K., Espinosa-Artilles, Gunatilaka, A.A.L., Stock, S.P., and Molnar, I.: Biosynthesis of the cyclooligomer depsipeptide bassianolide, an insecticidal virulence factor of Beauveria bassiana. Fungal Genet. Biol. 2009; 46: 353-364.
3. Xu, Y., Wijeratne, E.M.K., Espinosa-Artiles, P., Gunatilaka, A.A.L., and Molnar, I.: Combinatorial mutasynthesis of scrambled beauvericins, cyclooligomer depsipeptide cell migration inhibitors from Beauveria bassiana. ChemBioChem, 2009; 10: 345-354.
4. Wang, S., Xu, Y., Maine, E.A., Wijeratne, E.M.K., Espinosa-Artilles, P., Gunatilaka, A.A.L., and Molnar, I.: Functional characterization of the biosynthesis of radicicol, an Hsp90 inhibitor resorcylic acid lactone from Chaetomium chiversii. Chem. Biol. 2008; 15: 1328-1338.
5. Xu, Y., Orozco, R., Wijeratne, E.M.K., Gunatilaka, A.A.L., Stock, S.P., and Molnar, I.: Biosynthesis of the cyclooligomer depsipeptide beauvericin, a virulence factor of the entomopathogenic fungus Beauveria bassiana. Chem. Biol. 2008; 15: 898-907.
6. Xu, Y., Zhan, Z., Wijeratne, E.M.K., Burns, A.M., Gunatilaka, A.A.L. and Molnar, I.: Cytotoxic and anti-haptotactic beauvericin analogs from precursor-directed biosynthesis with the insect pathogen Beauveria bassiana ATCC 7159. J. Nat. Prod. 2007; 70: 1467-1471.
7. Trefzer, A., Jungmann, V., Molnar, I., Botejue, A., Buckel, D., Frey, G., Hill, D.S., Jorg, M., Ligon, J.M., Mason, D., Moore, D., Pachlatko, J.P., Richardson, T.H., Spangenberg, P., Wall, M.A., Zirkle, R., and Stege, J.T.: Biocatalytic conversion of avermectin to 4”-oxo-avermectin: Improvement of cytochrome P450 monooxygenase specificity by directed evolution. Appl. Env. Microbiol. 2007; 73: 4317-4325.
8. Molnar, I., Jungmann, V., Stege, J., Trefzer, A., and Pachlatko, J.P.: Biocatalytic conversion of avermectin into 4”-oxo-avermectin: discovery, characterization, heterologous expression and specificity improvement of the cytochrome P450 enzyme. Biochem. Soc. Trans. 2006; 34: 1236-1240.
9. Molnar, I., Hill, D.S., Zirkle, R., Hammer, P.E., Gross, F., Buckel, T.G., Jungmann, V., Pachlatko, J.P., and Ligon, J.M.: Biocatalytic conversion of avermectin to 4”-oxo-avermectin: heterologous expression of the ema1 cytochrome P450 monooxygenase. Appl. Environ. Microbiol. 2005; 71: 6977-6985.
10. Jungmann, V., Molnar, I., Hammer, P.E., Hill, D.S., Zirkle, R., Buckel, T.G., Buckel, D., Ligon, J.M., and Pachlatko, J.P.: Biocatalytic conversion of avermectin to 4”-oxo-avermectin: Characterization of biocatalytically active bacterial strains and of cytochrome P450 monooxygenase enzymes and their genes. Appl. Environ. Microbiol. 2005; 71: 6968-6976.
11. Zirkle, R., Ligon, J.M., Molnar, I.: Heterologous production of the antifungal polyketide antibiotic soraphen A of Sorangium cellulosum So ce26 in Streptomyces lividans. Microbiology 2004; 150: 2761-2774.
12. Zirkle, R., Ligon, J.M., Molnar, I.: Cloning, sequence analysis and disruption of the mglA gene involved in swarming motility of Sorangium cellulosum So ce26, a producer of the antifungal polyketide antibiotic soraphen A. J. Biosci. Bioeng. 2004; 97: 267-274.
13. Zirkle, R., Black, T.A., Gorlach, J., Ligon, J.M,, Molnar, I.: Analysis of a 108-kb region of the Saccharopolyspora spinosa genome covering the obscurin polyketide synthase locus. DNA Sequence 2004; 15: 123-134.
14. Nowak-Thompson, B,. Hammer, P.E., Hill, D.S., Stafford, J., Torkewitz, N.,Gaffney, T.D., Lam, S.T., Molnar, I., Ligon, J.M.: 2,5-dialkylresorcinol biosynthesis in Pseudomonas aurantiaca: a novel head-to-head condensation of two fatty acid-derived precursors. J. Bacteriol. 2003; 185: 860-869.
15. Ligon, J.M., Hill, S., Beck, J., Zirkle, R., Molnar, I., Zawodny, J., Money, S., Schupp, T.: Characterization of the biosynthetic gene cluster for the antifungal polyketide soraphen A from Sorangium cellulosum So ce26. Gene 2002; 285: 257-267.
16. Molnar, I., Schupp, T., Ono, M., Zirkle, R.E., Milnamow, M., Nowak-Thompson, B., Engel, N., Toupet, C., Stratmann, A., Cyr, D.D., Gorlach, J., Mayo, J.M., Hu, A., Goff, S., Schmid, J., Ligon, J.M.: The biosynthetic gene cluster for the microtubule-stabilizing agents epothilones A and B from Sorangium cellulosum So ce90. Chem. Biol. 2000; 7: 97-109.
17. König, A., Schwecke, T., Molnar, I., Böhm, G.A., Lowden, P.A.S., Staunton, J., Leadlay, P.F.: The pipecolate incorporating enzyme for the biosynthesis of the immunosuppressant rapamycin: Nucleotide sequence analysis, disruption, and heterologous expression of rapP from Streptomyces hygroscopicus. Eur. J. Biochem. 1997; 247: 526-534.
18. Molnar, I., Aparicio, J.F., Haydock, S.F., Khaw, L.E., Schwecke, T., König, A., Staunton, J., and Leadlay, P.F.: Organization of the biosynthetic gene cluster for rapamycin in Streptomyces hygroscopicus: Analysis of genes flanking the polyketide synthase. Gene 1996; 169: 9-16.
19. Aparicio, J.F., Molnar, I., Schwecke, T., König, A., Haydock, S.F., Khaw, L.E., Staunton, J., Leadlay, P.F.: Organization of the biosynthetic gene cluster for rapamycin in Streptomyces hygroscopicus: Analysis of the enzymatic domains in the modular polyketide synthase. Gene 1996; 169: 1-7.
20. Murooka, Y., Choi, K.-P., Molnar, I., Dziadek, J., Cho, H.-J., Nomura, N., Yamashita, M.: Recent advances in studies on Streptomyces cholesterol oxidase and gene clusters involved in steroid catabolism in prokaryotes. Actinomycetology 1996; 10: 1-11.
21. Haydock, S.F., Aparicio, J.F., Molnar, I., Schwecke, T., Khaw, L.E., König, A., Marsden, A.F.A., Galloway, I.S., Staunton, J., Leadlay, P.F.: Divergent sequence motifs correlated with the substrate specificity of (methyl)malonyl-CoA:acyl carrier protein transacylase domains in modular polyketide synthases. FEBS Letters 1995; 374: 246-248.
22. Schwecke, T., Aparicio, J.F., Molnar, I., König, A., Khaw, L.E., Haydock, S.F., Oliynyk, M., Caffrey, P., Cortés, J., Lester, J.B., Böhm, G.A., Staunton, J., Leadlay, P.F.: The biosynthetic gene cluster for the polyketide immunosuppressant rapamycin. Proc. Natl. Acad. Sci. USA 1995; 92: 7839-7843.
23. Choi, K.-P., Molnar, I., Yamashita, M., Murooka, Y.: Purification and characterisation of the 3-ketosteroid-Δ1-dehydrogenase of Arthrobacter simplex produced in Streptomyces lividans. J. Biochem. 1995; 117: 1043-1049.
24. Choi, K.-P., Molnar, I., Murooka, Y.: Secretory overproduction of Arthrobacter simplex 3-ketosteroid-Δ1-dehydrogenase by Streptomyces lividans with a multi-copy shuttle vector. Appl. Microbiol. Biotechnol. 1995; 43: 1044-1049.
25. Molnar, I., Choi, K.-P., Yamashita, M., Murooka, Y.: Molecular cloning, expression in Streptomyces lividans, and analysis of a gene cluster from Arthrobacter simplex encoding 3-ketosteroid-Δ1-dehydrogenase, 3-ketosteroid-Δ5-isomerase and a hypothetical regulatory protein. Mol. Microbiol. 1995; 15: 895-905.
26. Molnar, I., and Murooka, Y.: Helix-turn-helix DNA-binding motifs of Streptomyces - a cautionary note. Mol. Microbiol. 1993; 8: 783-784.
27. Molnar, I., and Murooka, Y.: Nucleotide sequence analysis of a region upstream of the cholesterol oxidase - cytochrome P450 operon of Streptomyces sp. SA-COO reveals repeating units coding for putative transmembrane and DNA-binding proteins. J. Ferment. Bioeng. 1993; 76: 257-264.
28. Molnar, I., Hayashi, N., Choi, K.-P., Yamamoto, H., Yamashita, M., and Murooka, Y.: Bacterial cholesterol oxidases are able to act as flavoprotein-linked ketosteroid monooxygenases that catalyse the hydroxylation of cholesterol to 4-cholesten-6-ol-3-one. Mol. Microbiol. 1993; 7: 419-428.
29. Molnar, I., Choi, K.-P., Hayashi, N., and Murooka, Y.: Secretory overproduction of Streptomyces cholesterol oxidase by Streptomyces lividans with a multi-copy shuttle vector. J. Ferment. Bioeng. 1991; 72: 368-372.
Chapters in scholarly books and monographs
1. Molnar, I., Zirkle, R., Ligon, J.M.: Biosynthesis of the antifungal polyketide soraphen A in Sorangium cellulosum and Streptomyces lividans. In: Polyketides: Biosynthesis, biological activity and genetic engineering. Baerson, S.R. and Rimando, A.M. (eds.), ACS Books #955, Washington, D.C., 2007: pp. 217-230. (ISBN10: 0-8412-3978-9).
2. Molnar, I.: Secretory overproduction of homologous and heterologous proteins by recombinant Streptomyces - What has been accomplished? In: Recombinant microbes for industrial and agricultural applications. Murooka, Y., and Imanaka, T. (eds.), Marcel Dekker, Inc., New York, 1994: pp. 81-103.
3. Murooka, Y., Yamashita, M., Takizawa, N., Molnar, I., Choi, K.-P.: Secretory overproduction of microbial enzymes using host-vector systems. In: Microbial utilization of renewable resources. Kinoshita, S., and Bhumiratana, A. (eds.), Osaka University Press, Osaka, 1991: pp. 219-227.
4. Molnar, I., Ambrus, G., Ott, I.: Transformation with shuttle DNA cloning vectors. In: Biotechnology of the Present. Szentirmai, A. (ed.), OMIKK, Budapest 1988.
Patents
1. Molnar, I., Ligon, J.M., Zirkle, R.E., Hammer, P.E., Hill, D.S., Pachlatko, J.P., Buckel, T.G.: WO 0292801; November 21, 2002.
2. Schupp, T., Ligon, J.M., Molnar, I., Zirkle, R., Cyr, D., Görlach, J.: US 6,383,787; May 7, 2002.
3. Schupp, T., Ligon, J.M., Molnar, I., Zirkle, R., Cyr, D., Görlach, J.: US 6,358,719; March 19, 2002.
4. Schupp, T., Ligon, J.M., Molnar, I., Zirkle, R., Cyr, D., Görlach, J.: US 6,355,459; March 12, 2002.
5. Schupp, T., Ligon, J.M., Molnar, I., Zirkle, R., Cyr, D., Görlach, J.: US 6,355,458; March 12, 2002.
6. Schupp, T., Ligon, J.M., Molnar, I., Zirkle, R., Cyr, D., Görlach, J.: US 6,355,457; March 12, 2002.
7. Schupp, T., Ligon, J.M., Molnar, I., Zirkle, R., Cyr, D., Görlach, J.: US 6,121,029; September 19, 2000.
8. Schupp, T., Ligon, J.M., Molnar, I., Zirkle, R., Cyr, D., Görlach, J.: US 6,346,404; February 12, 2002.
9. Ott, I., Kiss, G.B., Molnar, I., Kiss, P., Klupp, T., Szeleczky, Z., Ambrus, G.: EP 105848.1; US 08/044.506, art unit 1814; 1993.
10. Kiss, G.B., Vincze, E., Ott, I., Kiss, P., Klupp, T., Molnar, I., Szeleczky, Z., Ambrus, G., Moravcsik, I.: EP 308883; 1989.
11. Kiss, G.B., Vincze, E., Ott, I., Kiss, P., Klupp, T., Molnar, I., Szeleczky, Z., Ambrus, G., Moravcsik, I.: EP 115460.3; 1988.
