Shelley D. Copley
My lab studies the molecular evolution of enzymes and metabolic pathways. Metabolic enzymes are superb catalysts, accelerating chemical reactions by up to 20 orders of magnitude. Many, and probably all, also catalyze adventitious secondary reactions due to the highly reactive nature of their active sites. Although these “promiscuous” activities are often inefficient, they provide a repertoire of activities that can be drawn upon if conditions change and catalysis of a secondary reaction becomes important for fitness. Since most bacteria have 1000-2000 enzymes, and each enzyme probably has several promiscuous activities, bacteria can access many thousands of catalytic activities when a new enzyme is needed to enhance fitness or even to enable survival.
Evolution of a New Enzyme by Gene Duplication and Divergence
New enzymes often evolve from promiscuous activities of previously existing enzymes by a process of gene duplication/amplification and divergence. Previous studies of this process have focused solely on mutations in the gene undergoing duplication /amplification and divergence. We are taking a genome-wide approach to studying all of the mutations that contribute to fitness in the face of a novel evolutionary challenge.
ArgC (N-acetyl-L-g-glutamylphosphate reductase) is essential for growth of E. coli on glucose because it is required for arginine synthesis. However, a point mutation in proA allows ProA (glutamylphosphate reductase) to serve both functions, albeit poorly. We refer to this enzyme as ProA*. ProA* is the “weak-link” in metabolism; hence, cells are under strong selective pressure to increase its catalytic activity. We evolved eight lineages of a strain lacking ArgC and carrying ProA* under selection for improved synthesis of arginine (Figure 1). As expected, the region surrounding proA* amplifies to several copies. A mutation in proA* was detected only in population 3. In the other populations, we observed genomic mutations that increase arginine synthesis by either pushing or pulling material through the compromised biosynthetic pathway. This work demonstrates that evolution of a new enzyme by gene duplication and divergence is inextricably intertwined with genomic mutations that improve fitness by other mechanisms.
Promiscuity, Serendipity and Metabolic Innovation
Promiscuous enzymes can serve as the starting point for evolution of new enzyme activities. However, their evolutionary potential goes further; multiple promiscuous activities can be patched together to generate novel metabolic pathways. We call these pathways “serendipitous” because their assembly relies upon the fortuitous combination of available promiscuous activities, which can differ substantially in different organisms, or in the same organism under different environmental conditions.
We are studying the emergence of serendipitous pathways in a model system in which we have deleted a gene (pdxB) that is required for synthesis of the essential cofactor pyridoxal 5-phosphate (PLP, a.k.a. vitamin B6). We have succeeded in restoring robust growth of ∆pdxB E. coli within as few as 150 generations by a novel four-step serendipitous pathway. Each step of the serendipitous pathway is catalyzed by a promiscuous enzyme that normally serves another function. This work demonstrates the potential for evolution of novel metabolic pathways that resides within the proteome of a bacterium.
We discovered that a strikingly different suite of mutations was found when the same strain was evolved under different environmental conditions, suggesting that either a different solution to the evolutionary challenge was found, or that the same solution emerged but by a different mechanism. Current work addresses how PLP synthesis can be restored in strains evolved under other environmental conditions.
- Copley SD; Babbs S; Losoff B. (Jan 2021). Science and Society: Integrating Historical Science Materials Into an Undergraduate Biology Course. , 8. 10.24918/cs.2021.23
- Copley SD. (Sep 2020). The physical basis and practical consequences of biological promiscuity. Physical Biology , 17(5). 10.1088/1478-3975/ab8697
- Choudhury A; Fankhauser RG; Freed EF; Oh EJ; Morgenthaler AB; Bassalo MC; Copley SD; Kaar JL; Gill RT. (May 2020). Determinants for Efficient Editing with Cas9-Mediated Recombineering in Escherichia coli. ACS Synthetic Biology , 9(5), 1083-1099. 10.1021/acssynbio.9b00440
- Copley SD. (Apr 2020). Evolution of new enzymes by gene duplication and divergence. The Federation of European Biochemical Societies (FEBS) Journal , 287(7), 1262-1283. 10.1111/febs.15299
- Morgenthaler, AB; Kinney, WR; Ebmeier, CC; Walsh, CM; Snyder, DJ; Cooper, VS; Old, WM; Copley, SD. (9-Dec 2019). Mutations that improve efficiency of a weak-link enzyme are rare compared to adaptive mutations elsewhere in the genome. ELIFE, 8. 10.7554/eLife.53535
- Kim, J; Flood, JJ; Kristofich, MR; Gidfar, C; Morgenthaler, AB; Fuhrer, T; Sauer, U; Snyder, D; Cooper, VS; Ebmeier, CC; Old, WM; Copley, SD. (26-Nov 2019). Hidden resources in the Escherichia coli genome restore PLP synthesis and robust growth after deletion of the essential gene pdxB. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 116(48), 24164-24173. 10.1073/pnas.1915569116
- Flood, JJ; Copley, SD. (NOV-DEC 2018). Genome-Wide Analysis of Transcriptional Changes and Genes That Contribute to Fitness during Degradation of the Anthropogenic Pollutant Pentachlorophenol by Sphingobium chlorophenolicum. MSYSTEMS, 3(6). 10.1128/mSystems.00275-18
- Kristofich, J, AB Morgenthaler, WR Kinney, CC Ebmeier, DJ Snyder, WM Old, VS Cooper and SD Copley. (AUG 2018). Synonymous mutations make dramatic contributions to fitness when growth is limited by a weak-link enzyme. PLOS GENETICS, 14(8). 10.1371/journal.pgen.1007615
- Mikkonen, A, K Ylaranta, M Tiirola, LAL Dutra, P Salmi, M Romantschuk, S Copley, J Ikaheimo and A Sinkkonen. (1-Jul 2018). Successful aerobic bioremediation of groundwater contaminated with higher chlorinated phenols by indigenous degrader bacteria. WATER RESEARCH, 138, 118-128. 10.1016/j.watres.2018.03.033
- Copley, SD. (DEC 2017). Shining a light on enzyme promiscuity. CURRENT OPINION IN STRUCTURAL BIOLOGY, 47, 167-175. 10.1016/j.sbi.2017.11.001
- Kershner, JP, SY McLoughlin, J Kim, A Morgenthaler, CC Ebmeier, WM Old and SD Copley. (OCT 2016). A Synonymous Mutation Upstream of the Gene Encoding a Weak-Link Enzyme Causes an Ultrasensitive Response in Growth Rate. JOURNAL OF BACTERIOLOGY, 198(20), 2853-2863. 10.1128/JB.00262-16
- Thiaville, JJ, J Flood, S Yurgel, L Prunetti, M Elbadawi-Sidhu, G Hutinet, F Forouhar, XS Zhang, V Ganesan, P Reddy, O Fiehn, JA Gerlt, JF Hunt, SD Copley and V de Crecy-Lagard. (AUG 2016). Members of a Novel Kinase Family (DUF1537) Can Recycle Toxic Intermediates into an Essential Metabolite. ACS CHEMICAL BIOLOGY, 11(8), 2304-2311. 10.1021/acschembio.6b00279
- Khanal, A, SY McLoughlin, JP Kershner and SD Copley. (JAN 2015). Differential Effects of a Mutation on the Normal and Promiscuous Activities of Orthologs: Implications for Natural and Directed Evolution. MOLECULAR BIOLOGY AND EVOLUTION, 32(1), 100-108. 10.1093/molbev/msu271
- Copley, SD. (FEB 2015). An evolutionary biochemists perspective on promiscuity. TRENDS IN BIOCHEMICAL SCIENCES, 40(2), 72-78. 10.1016/j.tibs.2014.12.004
- Copley, SD. (DEC 2014). An evolutionary perspective on protein moonlighting. BIOCHEMICAL SOCIETY TRANSACTIONS, 42, 1684-1691. 10.1042/BST20140245
- Kim, J, AM Webb, JP Kershner, S Blaskowski and SD Copley. (25-Sep 2014). A versatile and highly efficient method for scarless genome editing in Escherichia coli and Salmonella enterica. BMC BIOTECHNOLOGY, 14. 10.1186/1472-6750-14-84
- Rudolph, J, AH Erbse, LS Behlen and SD Copley. (21-Oct 2014). A Radical Intermediate in the Conversion of Pentachlorophenol to Tetrachlorohydroquinone by Sphingobium chlorophenolicum. BIOCHEMISTRY, 53(41), 6539-6549. 10.1021/bi5010427
- Rokicki, J, D Knox, RD Dowell and SD Copley. (24-Jan 2014). CodaChrome: a tool for the visualization of proteome conservation across all fully sequenced bacterial genomes. BMC GENOMICS, 15. 10.1186/1471-2164-15-65
- Novikov, Y and SD Copley. (13-Aug 2013). Reactivity landscape of pyruvate under simulated hydrothermal vent conditions. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 110(33), 13283-13288. 10.1073/pnas.1304923110
- Yadid, I, J Rudolph, K Hlouchova and SD Copley. (11-Jun 2013). Sequestration of a highly reactive intermediate in an evolving pathway for degradation of pentachlorophenol. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 110(24), E2182-E2190. 10.1073/pnas.1214052110
- Kim, J and SD Copley. (30-Apr 2013). The Orphan Protein Bis-gamma-glutamylcystine Reductase Joins the Pyridine Nucleotide Disulfide Reductase Family. BIOCHEMISTRY, 52(17), 2905-2913. 10.1021/bi4003343
- Kim, J and SD Copley. (16-Oct 2012). Inhibitory cross-talk upon introduction of a new metabolic pathway into an existing metabolic network. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 109(42), E2856-E2864. 10.1073/pnas.1208509109
- Copley, SD. (JUL 2012). Moonlighting is mainstream: Paradigm adjustment required. BIOESSAYS, 34(7), 578-588. 10.1002/bies.201100191
- Hlouchova, K, J Rudolph, JMH Pietari, LS Behlen and SD Copley. (8-May 2012). Pentachlorophenol Hydroxylase, a Poorly Functioning Enzyme Required for Degradation of Pentachlorophenol by Sphingobium chlorophenolicum. BIOCHEMISTRY, 51(18), 3848-3860. 10.1021/bi300261p
- Copley, SD, J Rokicki, P Turner, H Daligault, M Nolan and M Land. (2012). The Whole Genome Sequence of Sphingobium chlorophenolicum L-1: Insights into the Evolution of the Pentachlorophenol Degradation Pathway. GENOME BIOLOGY AND EVOLUTION, 4(2), 184-198. 10.1093/gbe/evr137
- Copley, SD. (2-Jan 2012). Toward a Systems Biology Perspective on Enzyme Evolution. JOURNAL OF BIOLOGICAL CHEMISTRY, 287(1), 3-10. 10.1074/jbc.R111.254714
- Chumachenko, N, Y Novikov, RK Shoemaker and SD Copley. (18-Nov 2011). A Dimethyl Ketal-Protected Benzoin-Based Linker Suitable for Photolytic Release of Unprotected Peptides. JOURNAL OF ORGANIC CHEMISTRY, 76(22), 9409-9416. 10.1021/jo2017263
- Novikov, Y, SD Copley and BE Eaton. (20-Apr 2011). A simple route for synthesis of 4-phospho-D-erythronate. TETRAHEDRON LETTERS, 52(16), 1913-1915. 10.1016/j.tetlet.2011.02.045
- Copley, SD. (2010). Evolution and the Enzyme. COMPREHENSIVE NATURAL PRODUCTS II: CHEMISTRY AND BIOLOGY, VOL 8: ENZYMES AND ENZYME MECHANISMS, 9-46.
- Kim, JH, JP Kershner, Y Novikov, RK Shoemaker and SD Copley. (NOV 2010). Three serendipitous pathways in E. coli can bypass a block in pyridoxal-5 -phosphate synthesis. MOLECULAR SYSTEMS BIOLOGY, 6. 10.1038/msb.2010.88
- Rudolph, J, J Kim and SD Copley. (2-Nov 2010). Multiple Turnovers of the Nicotino-Enzyme PdxB Require alpha-Keto Acids as Cosubstrates. BIOCHEMISTRY, 49(43), 9249-9255. 10.1021/bi101291d
- Copley, SD. (AUG 2009). Evolution of efficient pathways for degradation of anthropogenic chemicals. NATURE CHEMICAL BIOLOGY, 5(8), 560-567. 10.1038/nchembio.197
- Hamady, M, J Widmann, SD Copley and R Knight. (2008). MotifCluster: an interactive online tool for clustering and visualizing sequences using shared motifs. GENOME BIOLOGY, 9(8). 10.1186/gb-2008-9-8-r128
- McLoughlin, SY and SD Copley. (9-Sep 2008). A compromise required by gene sharing enables survival: Implications for evolution of new enzyme activities. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 105(36), 13497-13502. 10.1073/pnas.0804804105
- Warner, JR, LS Behlen and SD Copley. (11-Mar 2008). A trade-off between catalytic power and substrate inhibition in TCHQ dehalogenase. BIOCHEMISTRY, 47(10), 3258-3265. 10.1021/bi702431n
- Copley, SD, E Smith and HJ Morowitz. (DEC 2007). The origin of the RNA world: Co-evolution of genes and metabolism. BIOORGANIC CHEMISTRY, 35(6), 430-443. 10.1016/j.bioorg.2007.08.001
- Kim, J and SD Copley. (6-Nov 2007). Why metabolic enzymes are essential or nonessential for growth of Escherichia coli k12 on glucose. BIOCHEMISTRY, 46(44), 12501-12511. 10.1021/bi7014629
- Warner, JR and SD Copley. (13-Nov 2007). Pre-steady-state kinetic studies of the reductive dehalogenation catalyzed by tetrachlorohydroquinone dehalogenase. BIOCHEMISTRY, 46(45), 13211-13222. 10.1021/bi701069n
- Warner, JR and SD Copley. (10-Apr 2007). Mechanism of the severe inhibition of tetrachlorohydroquinone dehalogenase by its aromatic substrates. BIOCHEMISTRY, 46(14), 4438-4447. 10.1021/bi0620104
- Copley, SD, E Smith and HJ Morowitz. (22-Mar 2005). A mechanism for the association of amino acids with their codons and the origin of the genetic code. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 102(12), 4442-4447. 10.1073/pnas.0501049102
- Warner, JR, SL Lawson and SD Copley. (2-Aug 2005). A mechanistic investigation of the thiol-disulfide exchange step in the reductive dehalogenation catalyzed by tetrachlorohydroquinone dehalogenase. BIOCHEMISTRY, 44(30), 10360-10368. 10.1021/bi050666b
- Dai, MH, S Ziesman, T Ratcliffe, RT Gill and SD Copley. (JAN 2005). Visualization of protoplast fusion and quantitation of recombination in fused protoplasts of auxotrophic strains of Escherichia coli. METABOLIC ENGINEERING, 7(1), 45-52. 10.1016/j.ymben.2004.09.002
- Cleland, CE and SD Copley. (OCT 2005). The possibility of alternative microbial life on Earth. INTERNATIONAL JOURNAL OF ASTROBIOLOGY, 4(4-Mar), 165-173. 10.1017/S147355040500279X
- Copley, SD, WRP Novak and PC Babbitt. (9-Nov 2004). Divergence of function in the thioredoxin fold suprafamily: Evidence for evolution of peroxiredoxins from a thioredoxin-like ancestor. BIOCHEMISTRY, 43(44), 13981-13995. 10.1021/bi048947r
- Dai, MH and SD Copley. (APR 2004). Genome shuffling improves degradation of the anthropogenic pesticide pentachlorophenol by Sphingobium chlorophenolicum ATCC 39723. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 70(4), 2391-2397. 10.1128/AEM.70.4.2391-2397.2004
- Dai, MH, JB Rogers, JR Warner and SD Copley. (JAN 2003). A previously unrecognized step in pentachlorophenol degradation in Sphingobium chlorophenolicum is catalyzed by tetrachlorobenzoquinone reductase (PcpD). JOURNAL OF BACTERIOLOGY, 185(1), 302-310. 10.1128/JB.185.1.302-310.2003
- Copley, SD. (APR 2003). Enzymes with extra talents: moonlighting functions and catalytic promiscuity. CURRENT OPINION IN CHEMICAL BIOLOGY, 7(2), 265-272. 10.1016/S1367-5931(03)00032-2
- Kiefer, PM and SD Copley. (29-Jan 2002). Characterization of the initial steps in the reductive dehalogenation catalyzed by tetrachlorohydroquinone dehalogenase. BIOCHEMISTRY, 41(4), 1315-1322. 10.1021/bi0117504
- Kiefer, PM, DL McCarthy and SD Copley. (29-Jan 2002). The reaction catalyzed by tetrachlorohydroquinone dehalogenase does not involve nucleophilic aromatic substitution. BIOCHEMISTRY, 41(4), 1308-1314. 10.1021/bi0117495
- Copley, SD, K Anandarajah and PJ Kiefer. (28-Feb 2001). A tale of two enzymes: tetrachlorohydroquinone dehalogenase and maleylacetoacetate isomerase. CHEMICO-BIOLOGICAL INTERACTIONS, 133(3-Jan), 200-203.
- Copley, SD. (JUN 2000). Evolution of a metabolic pathway for degradation of a toxic xenobiotic: the patchwork approach. TRENDS IN BIOCHEMICAL SCIENCES, 25(6), 261-265. 10.1016/S0968-0004(00)01562-0
- Fall, R and SD Copley. (APR 2000). Bacterial sources and sinks of isoprene, a reactive atmospheric hydrocarbon. ENVIRONMENTAL MICROBIOLOGY, 2(2), 123-130. 10.1046/j.1462-2920.2000.00095.x
- Anandarajah, K, PM Kiefer, BS Donohoe and SD Copley. (9-May 2000). Recruitment of a double bond isomerase to serve as a reductive dehalogenase during biodegradation of pentachlorophenol. BIOCHEMISTRY, 39(18), 5303-5311. 10.1021/bi9923813
- Xu, L, K Resing, SL Lawson, PC Babbitt and SD Copley. (15-Jun 1999). Evidence that pcpA encodes 2,6-dichlorohydroquinone dioxygenase, the ring cleavage enzyme required for pentachlorophenol degradation in Sphingomonas chlorophenolica strain ATCC 39723. BIOCHEMISTRY, 38(24), 7659-7669. 10.1021/bi990103y
- Copley, SD. (OCT 1998). Microbial dehalogenases: enzymes recruited to convert xenobiotic substrates. CURRENT OPINION IN CHEMICAL BIOLOGY, 2(5), 613-617. 10.1016/S1367-5931(98)80092-6
- Copley, SD. (MAR 1997). Diverse mechanistic approaches to difficult chemical transformations: Microbial dehalogenation of chlorinated aromatic compounds. CHEMISTRY & BIOLOGY, 4(3), 169-174. 10.1016/S1074-5521(97)90285-4
- McCarthy, DL, DF Louie and SD Copley. (19-Nov 1997). Identification of a covalent intermediate between glutathione and cysteine13 formed during catalysis by tetrachlorohydroquinone dehalogenase. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 119(46), 11337-11338. 10.1021/ja9726365
- McCarthy, DL, AA Claude and SD Copley. (MAY 1997). In vivo levels of chlorinated hydroquinones in a pentachlorophenol-degrading bacterium. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 63(5), 1883-1888.
- Willett, WS and SD Copley. (OCT 1996). Identification and localization of a stable sulfenic acid in peroxide-treated tetrachlorohydroquinone dehalogenase using electrospray mass spectrometry. CHEMISTRY & BIOLOGY, 3(10), 851-857.
- McCarthy, DL, S Navarrete, WS Willett, PC Babbitt and SD Copley. (19-Nov 1996). Exploration of the relationship between tetrachlorohydroquinone dehalogenase and the glutathione S-transferase superfamily. BIOCHEMISTRY, 35(46), 14634-14642. 10.1021/bi961730f
- CROOKS, GP, L XU, RM BARKLEY and SD COPLEY. (8-Nov 1995). EXPLORATION OF POSSIBLE MECHANISMS FOR 4-CHLOROBENZOYL COA DEHALOGENASE - EVIDENCE FOR AN ARYL-ENZYME INTERMEDIATE. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 117(44), 10791-10798. 10.1021/ja00149a001
- CROOKS, GP and SD COPLEY. (27-Sep 1994). PURIFICATION AND CHARACTERIZATION OF 4-CHLOROBENZOYL COA DEHALOGENASE FROM ARTHROBACTER SP STRAIN 4-CB1. BIOCHEMISTRY, 33(38), 11645-11649. 10.1021/bi00204a028
- CROOKS, GP and SD COPLEY. (14-Jul 1993). A SURPRISING EFFECT OF LEAVING GROUP ON THE NUCLEOPHILIC AROMATIC-SUBSTITUTION REACTION CATALYZED BY 4-CHLOROBENZOYL-COA DEHALOGENASE. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 115(14), 6422-6423. 10.1021/ja00067a072
- COPLEY, SD, E FRANK, WM KIRSCH and TH KOCH. (14-Feb 1992). DETECTION AND POSSIBLE ORIGINS OF AMINOMALONIC ACID IN PROTEIN HYDROLYSATES. ANALYTICAL BIOCHEMISTRY, 201(1), 152-157. 10.1016/0003-2697(92)90188-D
- COPLEY, SD and GP CROOKS. (APR 1992). ENZYMATIC DEHALOGENATION OF 4-CHLOROBENZOYL COENZYME-A IN ACINETOBACTER SP STRAIN 4-CB1. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 58(4), 1385-1387.
- GUILFORD, WJ, SD COPLEY and JR KNOWLES. (5-Aug 1987). ON THE MECHANISM OF THE CHORISMATE MUTASE REACTION. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 109(16), 5013-5019. 10.1021/ja00250a041
- COPLEY, SD and JR KNOWLES. (5-Aug 1987). THE CONFORMATIONAL EQUILIBRIUM OF CHORISMATE IN SOLUTION - IMPLICATIONS FOR THE MECHANISM OF THE NONENZYMATIC AND THE ENZYME-CATALYZED REARRANGEMENT OF CHORISMATE TO PREPHENATE. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 109(16), 5008-5013. 10.1021/ja00250a040
- COPLEY, SD and JR KNOWLES. (1985). THE UNCATALYZED CLAISEN REARRANGEMENT OF CHORISMATE TO PREPHENATE PREFERS A TRANSITION-STATE OF CHAIRLIKE GEOMETRY. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 107(18), 5306-5308. 10.1021/ja00304a064
- GRIMSHAW, CE, SG SOGO, SD COPLEY and JR KNOWLES. (1984). SYNTHESIS OF STEREOSELECTIVELY LABELED [9-H-2,H-3] CHORISMATE AND THE STEREOCHEMICAL COURSE OF "5-ENOLPYRUVOYLSHIKIMATE-3-PHOSPHATE SYNTHETASE. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 106(9), 2699-2700. 10.1021/ja00321a037
- KOVACHY, RJ, SD COPLEY and RH ALLEN. (1983). RECOGNITION, ISOLATION, AND CHARACTERIZATION OF RAT-LIVER D-METHYLMALONYL COENZYME-A HYDROLASE. JOURNAL OF BIOLOGICAL CHEMISTRY, 258(18), 1415-1421.