Faculty Profile

Dr. Emmanuel S. Burgos, Ph.D.

Emmanuel S. Burgos, Ph.D.

Research Assistant Professor, Department of Biochemistry

Areas of Research: Nicotinamide recycling and targeted inhibition of NAMPT for the development of novel therapeutics. Chemoenzymatic synthesis of co-factors and HTS platforms to identify selective methyltransferases' inhibitors.

Professional Interests

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BREAKING NEWS Our research got published in Chemical Science. With our collaborators from the Department of Molecular Pharmacology, we developed a novel analytical platform for the detection of methyltransferase activity (validated for in vivo and in vitro). Request your kit of the 1-Step EZ-MTase assay TODAY, it is FREE! Read Chemical Science here.

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Using top-notch analytical platform, synthetic and enzyme-assisted chemistry, I decipher enzyme mechanisms. My research aims toward designing small molecule inhibitors of therapeutic targets. I am an expert in rational drug design and I have an extensive knowledge, both theoretical and practical, in Analytical Chemistry, Synthetic Chemistry, Enzymology and Biochemistry.

Nicotinamide within the NAD Metabolism

Nicotinamide phosphoribosyltransferase (NAMPT) catalyzes the first reversible step in NAD biosynthesis and nicotinamide (NAM) salvage (Figure 1A; red). The enzyme is designed for efficient capture of nicotinamide by energetic coupling of ATP-hydrolysis to assist in extraordinary NAM binding affinity[13,14] and formation of nicotinamide mononucleotide (NMN). NAMPT provides the mechanism to replenish the NAD pool in human metabolism. In addition to its role in RedOx biochemistry, NAD fuels the sirtuins (SIRTs) to regulate transcription factors involved in pathways linked to inflammation, diabetes and lifespan (Figure 1A; blue).

NAMPT-mediated lifespan expansion has caused a focus on the catalytic mechanism, regulation and inhibition of NAMPT. Structural, mechanistic and inhibitor design all contribute to a developing but yet incomplete story of NAMPT function. Although the first generation of NAMPT inhibitors has entered clinical trials, disappointing outcomes suggest more powerful and specific inhibitors will be needed.

Understanding the ATP-linked mechanism of NAMPT and the catalytic site machinery may permit the design of improved NAMPT inhibitors as more efficient drugs against cancer[11]. Design of such inhibitors may be achieved by rational design using the Kinetic Isotope Effect tool (KIE; Figure 1B-1C)[7].

Use of Biocatalysts for Efficient Synthesis of Valuable Chemical Probes

Enzymes are powerful catalysts. Unlike tedious multi-step chemical syntheses, enzyme-assisted reactions deliver 1) high yields of pure products and 2) molecules with the desired stereochemistry. Furthermore, this approach does not require toxic organic solvents (green chemistry). Thus, we took advantage of these efficient biocatalysts. We harvested the enzymatic activity of NAD-nucleosidase mutants for efficient gram-scale production of O-acetyl-ADP-ribose (OAADPR; Figure 2D). This molecule is a by-product of the sirtuin-catalyzed reactions. OAADPR was obtained through our in-line production (99% yield; Figure 2A). This set-up allows for easy and fast preparation of pure metabolite[5]. Likewise, kinases were used to generate a phosphorylated derivative of Immucillin-A (Figure 3). ImmA-TP is a potent inhibitor of the newly described C–P lyase nucleosidase PhnI[6].

Pursuing our efforts, we use a chemo-enzymatic approach to generate artificial co-factor scaffolds. Methyltransferases (MTases) consume S-adenosyl-L-methionine (SAM, AdoMet) to deposit methyl marks onto acceptor substrates (e.g. lysine, arginine, DNA, RNA). Structural biology describes the SAM binding mode for several MTase candidates; such structures support our hypothesis that edited co-factors may display an enhanced affinity for specific targets. The chemical diversity within our library (Figure 4A) is a first step towards the development of isozyme-specific protein methyltransferases’ inhibitors.

Development of in vitro Assays to Decipher Enzymes Mechanisms and Identify Novel Therapeutic

By screening a wide range of ligands in a 384-well plate format, the thermal shift assay (TSA; Figure 4B) easily identifies promising chemical scaffolds. To further quantify affinity parameters, we developed highly sensitive coupled assays for the detection of ATP and its derivatives (e.g. adenosine, adenine, S-adenosyl-L-homocysteine SAH) by Photinus pyralis luciferase. These platforms are well suited for the detection of various enzymatic activities, including slow enzymes such as MTases (Figure 5)[10]. Using this technology, we provided the first most quantitative analysis for the Protein Arginine MethylTransferase (PRMT5) reaction mechanism (Figure 6)[2].

To characterize methyltransferases in vitro and in vivo, we recently reported a highly-sensitive one-step deaminase-linked continuous assay where the S-adenosyl-L-homocysteine (SAH) enzyme-product is rapidly and quantitatively catabolized to S-inosyl-L-homocysteine (SIH; Figure 7)[1]. The coupling deaminase (TM0936) displays robust activity over a broad pH-range, thus supporting the broad capabilities of the 1-Step EZ-MTase platform. Unlike discontinuous radioactive- and antibody-based assays, our method provides a simple, versatile and affordable approach towards the characterization of methyltransferases.

With its three logs of linear dynamic range, the 1-Step EZ-MTase can accurately detect methylation rates as low as 2 uM h-1. Because this analytical tool box senses small variations in SAM consumption, the 1-Step EZ-MTase provides an unprecedented approach to probing the effect of histone tail modifications (e.g. lysine acetylation, serine phosphorylation; Figure 7) on lysine/arginine methyltransferases catalyzed reactions. This analysis may elucidate the complex interplays of histone modifications and promote our understanding of the Histone Code.

Selected Publications


[1Emmanuel S. Burgos, , and David Shechter. 2017. “A simplified characterization of S-adenosyl-L-methionine-consuming enzymes with 1-Step EZ-MTase: a universal and straightforward coupled-assay for in vitro and in vivo setting.” Chem. Sci., 8: 6601-12. doi: 10.1039/C7SC02830J

[2Emmanuel S. Burgos, Carola Wilczek, Takashi Onikubo, Jeffrey B. Bonanno, Janina Jansong, Ulf Reimer, and David Shechter. 2015. “Histone H2A and H4 N-terminal tails are positioned by the MEP50 WD repeat protein for efficient methylation by the PRMT5 arginine methyltransferase.” J. Biol. Chem., 290(15):9674-89. PMID: 25713080 doi: 10.1074/jbc.M115.636894

[3] Keisha Thomas, Scott A. Cameron, Steven C. Almo, Emmanuel S. Burgos, Shivali A. Gulab, and Vern L. Schramm. 2015. “Active site and remote contributions to catalysis in methylthioadenosine nucleosidases.” Biochemistry, 54(15):2520-29 PMID: 25806409 doi: 10.1021/bi501487w

[4] Takashi Onikubo, Joshua J. Nicklay, Li Xing, Christopher Warren, Brandon Anson, Wei-Lin Wang, Emmanuel S. Burgos, Sophie E. Ruff, Jeffrey Shabanowitz, R. Holland Cheng, Donald F. Hunt, and David Shechter. 2015. “Developmentally Regulated Post-translational Modification of Nucleoplasmin Controls Histone Sequestration and Deposition.” Cell Rep., 10(10): 1735-48. PMID: 25772360 doi: 10.1016/j.celrep.2015.02.038

[5] Brett M. Hirsch, Emmanuel S. Burgos, and Vern L. Schramm. 2014. “Transition-state analysis of 2-O-acetyl-ADP-ribose hydrolysis by human macrodomain 1.” ACS Chem. Biol., 9(10):2255-62. PMID: 25051211 doi: 10.1021/cb500485w

[6] Siddhesh S. Kamat, Emmanuel S. Burgos, and Frank M. Raushel. 2013. “Potent inhibition of the C–P lyase nucleosidase PhnI by Immucillin-A triphosphate.” Biochemistry, 52(42):7366-8. PMID: 24111876 doi: 10.1021/bi4013287

[7Emmanuel S. Burgos, Mathew J. Vetticatt , and Vern L. Schramm. 2013. “Recycling nicotinamide. The transition-state structure of human nicotinamide phosphoribosyltransferase.” J. Am. Chem. Soc., 135(9):3485-93. PMID: 23373462 doi: 10.1021/ja310180

[8] Shaun B. Reeksting, Ingrid B. Müller, Pieter B. Burger, Emmanuel S. Burgos, Laurent Salmon, Abraham I. Louw, Lyn-Marie Birkholtz, and Carsten Wrenger. 2013. “Exploring inhibition of Pdx1, a component of the PLP synthase complex of the human malaria parasite Plasmodium falciparum.” Biochem J., 449(1):175-87. PMID: 23039077 doi: 10.1042/BJ20120925

[9] Keisha Thomas, Antti M. Haapalainen, Emmanuel S. Burgos, Gary B. Evans, Peter C. Tyler, Shivali Gulab, Rong Guan, and Vern L. Schramm. 2012. “Femtomolar inhibitors bind to 5′-methylthioadenosine nucleosidases with favorable enthalpy and entropy.” Biochemistry, 51(38):7541-50. PMID: 22931458 doi: 10.1021/bi3009938

[10Emmanuel S. Burgos, Shivali A. Gulab, María B. Cassera, and Vern L. Schramm. 2012. “Luciferase-based assay for adenosine: application to S-adenosyl-L-homocysteine hydrolase.” Anal. Chem., 84(8):3593-98. PMID: 22416759 doi: 10.1021/ac203297z

[11Emmanuel S. Burgos. 2011. “NAMPT in regulated NAD biosynthesis and its pivotal role in human metabolism.” Curr. Med. Chem., 18(13):1947-61. PMID: 21517777

[12] María B. Cassera, Meng-Chiao Ho, Emilio F. Merino, Emmanuel S. Burgos, Agnes Rinaldo-Matthis, Steven C. Almo, and Vern L. Schramm. 2011. “A high-affinity adenosine kinase from Anopheles gambiae.” Biochemistry, 50(11):1885-93. PMID: 21247194 doi: 10.1021/bi101921w

[13Emmanuel S. Burgos, Meng-Chiao Ho, Steven C. Almo, and Vern L. Schramm. 2009. “A phosphoenzyme mimic, overlapping catalytic sites and reaction coordinate motion for human NAMPT.” Proc. Natl. Acad. Sci., 106(33):13748-53. PMID: 19666527 doi: 10.1073/pnas.0903898106

[14Emmanuel S. Burgos, and Vern L. Schramm. 2008. “Weak coupling of ATP hydrolysis to the chemical equilibrium of human nicotinamide phosphoribosyltransferase.” Biochemistry, 47(42):11086-96. PMID: 18823127 doi: 10.1021/bi801198m


Emmanuel S. Burgos, and Vern L. Schramm. August 1, 2013. “Luciferase-linked methods for detecting adenosine and uses thereof.” WO2013112358 A1.

Emmanuel S. Burgos, and David Shechter. “Chemoenzymatic synthesis of S-nucleosyl amino acids (SNA), analogs of S-adenosyl-L-methionine and S-adenosyl-L-homocysteine and uses thereof”, Filed February 3, 2016, U.S. Provisional Patent Application No. 62/290,502; Filed January 25, 2017, International Patent Application No. PTC/US2017/14804.

Emmanuel S. Burgos, and David Shechter. “Characterization of S-adenosyl-L-methionine consuming enzymes with 1-Step EZ-MTase: a universal coupled-assay”, Filed February 23, 2017, U.S. Provisional Patent Application No. 62/462,429.

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