Research Overview

Research in the Townsend Lab is driven by fundamental interests in natural products: their chemistry, biosynthesis, combinatorial and engineered biosynthesis, enzyme mechanism, drug design and synthesis to combat infectious diseases, cancer and obesity. The diversity of our research is reflected in the students who comprise the lab, coming from programs in Chemistry, Chemical Biology, Pharmacology, Biology, and Molecular Biophysics. Our lab has major strengths in the techniques of organic chemistry, molecular biology, mechanistic enzymology, microbiology and bioinformatics.

Iterative polyketide natural products

Polyketide natural products encompass a structurally and functionally diverse family of molecules. Our long-standing interest in the biosynthesis of aflatoxin, a potent environmental carcinogen, led to groundbreaking results on the function of the polyketide synthases (PKS) responsible for the 'programmed' synthesis and complex regioselective cyclization of linear precursors. Current efforts are focused on understanding the PKS enzymes involved in the biosynthesis of calicheamicin, cercosporin, aromatic polyketides and mycotoxins.

Recent Reports

~ Newman, A.G.; Vagstad, A.L.; Storm, P.A.; Townsend, C. A. “Systematic Domain Swaps of Iterative, Non-reducing Polyketide Synthases Provide a Mechanistic Understanding and Rationale For Catalytic Reprogramming” J. Am. Chem. Soc. 2014, 136, 7348-7362.

~ Belecki, K.; Townsend, C. A. “Biochemical Determination of Enzyme-Bound Metabolites: Preferential Accumulation of a Programmed Octaketide on the Enediyne Polyketide Synthase CalE8” J. Am. Chem. Soc. 2013, 135(38), 14339-14348.

~ Vagstad, A.L.; Newman, A.G.; Storm, P.A.; Belecki, K.; Crawford, J.M.; Townsend, C. A. “Combinatorial Domain Swaps Provide Insights into the Rules of Fungal Polyketide Synthase Programming and the Rational Synthesis of Non-native Aromatic Products,” Angew. Chem. Int. Ed. 2013, 52, 1718-1721.

~ Vagstad, A.L.; Hill, E.A.; Labonte, J.W.; Townsend, C. A. “Characterization of a Fungal Thioesterase Having Claisen Cyclase and Deacetylase Activities in Melanin Biosynthesis,” Chem. & Biol. 2012, 19, 1525-1534.

Papers of Significant Interest

~ Crawford, J. M.; Townsend, C. A. “New insights into the formation of fungal aromatic polyketides,” Nature Rev. Microbiol. 2010, 12, 879-889.

~ Crawford, J. M.; Korman, T. P.; Labonte, J. W.; Vagstad, A. L.; Hill, E. A.; Kamari-Bidkorpeh, O.; Tsai, S.-C.; Townsend, C. A. “Structural Basis for Biosynthetic Programming of Fungal Aromatic Polyketide Cyclization,” Nature 2009, 461, 1139-1143.

~ Udwary, D. W.; Merski, M.; Townsend, C. A. “A Method for Prediction of the Locations of Linker Regions Within Large Multifunctional Proteins, and Application to a Type I Polyketide Synthase,” J. Mol. Biol. 2002, 323, 585-598.

~ Minto, R. E.; Townsend, C. A. “Enzymology and Molecular Biology of Aflatoxin Biosynthesis,” Chem. Rev. 1997, 97, 2537-2555.

Biosynthesis of β-lactam antibiotics

Penicillins and cephalosporins are therapeutic mainstays in human medicine and whose industrial production has benefited from classic research into their biosynthesis. However, other clinically effective β-lactam drugs are related to natural products whose biosynthesis is less well understood. We have elucidated the biosynthetic pathway of the simple carbapenem and determined the function of key enzymes in the production of clavulinic acid, thienamycin, and nocardicin. We continue to study these natural products using a combination of synthetic chemistry, biochemistry, microbiology and bioinfromatics.

Recent Reports

~ Gaudelli, N.M.; Townsend, C. A. “Epimerization and substrate gating by a TE domain in β-lactam antibiotic biosynthesis” Nat. Chem. Biol. 2014, 10, 251-258.

~ Li, R; Lloyd, E.P.; Moshos, K.A.; Townsend, C. A. “Identification and Characterization of the Carbapenem MM 4550 and its Gene Cluster in Streptomyces argenteolus ATCC 11009” Chembiochem 2014, 15(2), 320-331.

~ Gaudelli, N.M.; Townsend, C. A. “Stereocontrolled Synthesis of Peptide Thioesters Containing Modified Seryl Residues as Probes of Antibiotic Biosynthesis” J. Org. Chem. Soc. 2013, 78(13), 6412-6426.

~ Phelan, R.M.; Townsend, C. A. “Mechanistic Insights into the Bifunctional Non-Heme Iron Oxygenase Carbapenem Synthase by Active Site Saturation Mutagenesis” J. Am. Chem. Soc. 2013, 135(20), 7496-7502.

Papers of Significant Interest

~ Bodner, M. J.; Phelan, R. M.; Townsend, C. A. “A Catalytic Asymmetric Route to Carbapenems,” Org. Lett. 2009, 11, 3606-3609.

~ Townsend, C. A. “New Reactions in Clavulanic Acid Biosynthesis,” Curr. Opin. Chem. Biol. 2002, 6, 583-589.

~ Li, R.; Stapon, A.; Blanchfield, J. T.; Townsend, C. A. “Three Unusual Reactions Mediate Carbapenem and Carbapenam Biosynthesis,” J. Am. Chem. Soc. 2000, 122, 9296-9297.

~ Bachmann, B. O.; Li, R.; Townsend, C. A. “β-Lactam Synthetase: A New Biosynthetic Enzyme,” Proc. Natl. Acad. Sci. 1998, 95, 9082-9086.

Metabolic and enzyme engineering

GFPandPT

The natural products we study are often related to chemicals that are of pharmaceutical or industrial interest. We have probed how biosynthetic enzymes involved in both polyketide and β-lactam biosynthesis may be altered to produce new compounds and how metabolic engineering may be used to make strains with desirable chemical properties.

Recent Reports

~ Vagstad, A.L.; Newman, A.G.; Storm, P.A.; Belecki, K.; Crawford, J.M.; Townsend, C. A. “Combinatorial Domain Swaps Provide Insights into the Rules of Fungal Polyketide Synthase Programming and the Rational Synthesis of Non-native Aromatic Products,” Angew. Chem. Int. Ed. 2013, 52, 1718-1721. Designated VIP: Very Important Paper

~ Labonte, J.W.; Kudo, F.; Freeman, M.F.; Raber, M.L.; Townsend, C. A. “Engineering the synthetic potential of β-lactam synthetase and the importance of catalytic loop dynamics,” MedChemComm 2012, 3, 960-966.

~ Phelan, R. M.; DiPardo, B. J.; Townsend, C. A. “A High-Throughput Screen for the Engineered Production of β-Lactam Antibiotics,” ACS Chem. Biol. 2012, 7, 835-840.

Papers of Significant Interest

~ Sokha, T.; Heins, R. A.; Phelan, R. M.; Greisler, J. M.; Townsend, C. A.; Ostermeier, M. “An Externally Tunable Bacterial Band-Pass Filter,” Proc. Natl. Acad. Sci. 2009, 106, 10135-10140.

~ Li, R.; Townsend, C. A. “Rational Strain Improvement for Enhanced Clavulanic Acid Production by Genetic Engineering of the Glycolytic Pathway in Streptomyces clavuligerus,” Metabol. Eng. 2006, 8, 240-252.

~ Muller, U.; van Assema, F.; Gunsior, M.; Orf, S.; Kremer, S.; Schipper, D.; Wagemans, A.; Townsend, C; Sonke, T.; Bovenberg, R.; Wubbolts, M. “Metabolic Engineering of the E. coli l-Phenylalanine pathway for the production of d-Phenylglycine (d-Phg),” Metabol. Eng. 2006, 8, 196-208.

Mechanistic Enzymology

GFPandPT

Biosynthetic enzymes often perform chemically difficult and mechanistically interesting reactions. Our discovery and study of β-lactam synthetases has led to a thorough understanding of their chemical mechanism and has shown how protein dynamics affect catalysis. We continue to investigate complex molecular phenomena catalyzed by biosynthetic enzymes such as regiospecific polyketide cyclization , protein autoproteolysis, and remarkably diverse chemical reactions coupled to thioesterase activity.

Recent Reports

~ Buller, A.R.; Freeman, M.F.; Schildbach, J.F; Townsend, C. A. “Exploring the role of conformational heterogeneity in cis-autoproteolytic activation of ThnT” Biochemistry 2014, 53(26), 4273-4281.

~ Labonte, J.W.; Kudo, F.; Freeman, M.F.; Raber, M.L.; Townsend, C. A. “Engineering the synthetic potential of β-lactam synthetase and the importance of catalytic loop dynamics,” MedChemComm 2012, 3, 960-966.

~ Buller, A. R.; Freeman, M. R.; Wright, N. T.; Schildbach J. F.; Townsend, C. A. “Insights into cis-autoproteolysis reveal a reactive state formed through conformational rearrangement,” Proc. Natl. Acad. Sci. 2012, 109, 2308-2313.

~ Vagstad, A. L.; Bumpus, S. B.; Belecki, K.; Kelleher, N.L.; Townsend, C. A. “Interrogation of Global Active Site Occupancy of a Fungal Iterative Polyketide Synthase Reveals Strategies for Maintaining Biosynthetic Fidelity,” J. Am. Chem. Soc. 2012, 134, 6865-6877.

Papers of Significant Interest

~ Buller, A.R.; Townsend, C. A. “Intrinsic Evolutionary Constraints on Protease Structure, Enzyme Acylation, and the Identity of the Catalytic Triad,” Proc. Natl. Acad. Sci. 2013, 110(8), E653-661.

~ Crawford, J. M.; Korman, T. P.; Labonte, J. W.; Vagstad, A. L.; Hill, E. A.; Kamari-Bidkorpeh, O.; Tsai, S.-C.; Townsend, C. A. “Structural Basis for Biosynthetic Programming of Fungal Aromatic Polyketide Cyclization,” Nature 2009, 461, 1139-1143.

~ Raber, M. L.; Arnett, S. O.; Townsend, C. A. “A Conserved Tyrosyl-Glutamyl Catalytic Dyad in Evolutionarily Linked Enzymes: Carbapenam Synthetase and β-Lactam Synthetase,” Biochemistry 2009, 48, 4959-4971.

~ Miller, M. T.; Bachmann, B. O.; Townsend, C. A.; Rosenzweig, A. C. “The Catalytic Cycle of β-Lactam Synthetase Observed by x-Ray Crystallographic Snapshots,” Proc. Natl. Acad. Sci. 2002, 99, 14752-14757.

~ Challis, G. L.; Ravel, J.; Townsend, C. A. “Predictive, Structure-Based Model of Amino Acid Recognition by Nonribosomal Peptide Synthetase Adenylation Domains,” Chemistry & Biology 2000, 7, 211-224.

Fatty Acid Synthase Inhibition

FAS

An extensive web of collaborative research has been developed over several years with scientists at the Johns Hopkins School of Medicine. The unifying theme of this effort has been the selective modulation (inhibition and allosteric stimulation) of enzymes involved in fatty acid metabolism as a therapeutic approach to the treatment of a variety of human cancers, tuberculosis, and a potential breakthrough therapy for obesity. Members of the lab are engaged in mechanism- and structure-based drug design and synthesis and perform SAR studies on key enzymes with the goal to produce more potent and selective inhibitors.

Recent Reports

~ Outlaw, V.K.; Wydysh, E.A.; Vadlamundi, A.; Medghalchi, S.M.; Townsend, C. A. “Design, Synthesis, and Evaluation of 4- and 5-Substituted o-(Octanesulfonamido)benzoic Acids as Inhibitors of Glycerol-3-Phosphate Acyltransferase” Med. Chem. Commun. 2014, 5, 826-830.

~ Kuhajda, F. P.; Aja, S.; Tu, Y.; Han, W. F.; Medghalchi, S. M.; El Meskini, R.; Landree, L. E.; Peterson, J. M.; Daniels, K.; Wong, K.; Wydysh, E. A.; Townsend, C. A.; Ronnett, G. V. “Pharmacological glycerol-3-phosphate acyltransferase inhibition decreases food intake and adiposity and increases insulin sensitivity in diet-induced obesity,” Am. J. Physiol-Reg. I. 2011, 301, 116-130.

~ Wydysh, E. A.; Vadlamudi, A.; Medghalchi, S. M.; Townsend, C. A. “Design, synthesis, and biological evaluation of conformationally constrained glycerol 3-phosphate acyltransferase inhibitors,” Bioorg. Med. Chem. 2010, 18, 6470-6479.

Papers of Significant Interest

~ Aja, S.; Landree, L. E.; Kleman, A. M.; Medghalchi, S. M.; Vadlamudi, A.; McFadden, J. M.; Aplasca, A.; Hyun, J.; Plummer, E.; Daniels, K.; Kemm, M.; Townsend, C. A.; Thupari, J. N.; Kuhajda, F. P.; Moran, T. H.; Ronnett, G. V. “Pharmacological Stimulation of Brain Carnitine Palmitoyl-Transferase-1 Decreases Food Intake and Body Weight,” Am. J. Physiol. Regul. Integr. Comp. Physiol. 2008, 294, R352-R361.

~ McFadden, J. M.; Medghalchi, S. M.; Thupari, J. N.; Pinn, M, L.; Vadlamudi, A.; Miller, K. I.; Kuhajda, F. P.; Townsend, C. A. “Application of a Flexible Synthesis of (5R)-Thiolactomycin to Develop New Inhibitors of Type I Fatty Acid Synthase,” J. Med. Chem. 2005, 48, 946-961.

~ Loftus, T. M.; Jaworsky D. E.; Frehywot, G. L.; Townsend, C. A.; Ronnett, G. V.; Lane, M. D.; Kuhajda, F. P. “Reduced Food Intake and Body Weight in Mice Treated with Fatty Acid Synthase Inhibitors,” Science 2000, 288, 2379-2381.

~ Kuhajda, F. P.; Pizer, E. S.; Li J. N.; Mani, N. S.; Frehywot, G. L.; Townsend, C. A. “Synthesis and Anti-tumor Activity of an Inhibitor of Fatty Acid Synthase,” Proc. Natl. Acad. Sci. 2000, 97, 3450-3454.

Maintained by: Douglas Cohen ~ Updated: 2014.09.05:16:03