Characterisation of an Escherichia coli line that completely lacks ribonucleotide reduction yields insights into the evolution of parasitism and endosymbiosis

Adeolu M, Gupta RS. 2014. A phylogenomic and molecular marker based proposal for the division of the genus

Borrelia into two genera: The emended genus Borrelia containing only the members of the relapsing fever

Borrelia, and the genus borreliella gen nov containing the members of the Lyme disease Borrelia (Borrelia

burgdorferi sensu lato complex). Antonie van Leeuwenhoek 105:1049–1072. DOI: https://doi.org/10.1007/​

s10482-014-0164-x, PMID: 24744012

Andersson JO. 2000. Evolutionary genomics: Is buchnera a bacterium or an organelle? Current Biology

10:R866–R868. DOI: https://doi.org/10.1016/s0960-9822(00)00816-2, PMID: 11114534

Barbour AG, Adeolu M, Gupta RS. 2017. Division of the genus borrelia into two genera (corresponding to lyme

disease and relapsing fever groups) reflects their genetic and phenotypic distinctiveness and will lead to a

better understanding of these two groups of microbes. International Journal of Systematic and Evolutionary

Microbiology 67:2058–2067. DOI: https://doi.org/10.1099/ijsem.0.001815, PMID: 28141502

Benner SA, Ellington AD, Tauer A. 1989. Modern metabolism as a palimpsest of the rna world. PNAS 86:7054–

7058. DOI: https://doi.org/10.1073/pnas.86.18.7054, PMID: 2476811

Blaesi EJ, Palowitch GM, Hu K, Kim AJ, Rose HR, Alapati R, Lougee MG, Kim HJ, Taguchi AT, Tan KO,

Laremore TN, Griffin RG, Krebs C, Matthews ML, Silakov A, Bollinger JM, Allen BD, Boal AK. 2018. Metal-­free

class ie ribonucleotide reductase from pathogens initiates catalysis with a tyrosine-­derived

dihydroxyphenylalanine radical. PNAS 115:10022–10027. DOI: https://doi.org/10.1073/pnas.1811993115,

PMID: 30224458

Bushnell B. 2014. BBTools. Software Package. http://bbtools.jgi.doe.gov

Cotruvo JA, Stubbe J. 2010. An active dimanganese (III) -tyrosyl radical cofactor in Escherichia coli class Ib

ribonucleotide reductase. Biochemistry 49:1297–1309. DOI: https://doi.org/10.1021/bi902106n, PMID:

20070127

Cotruvo JA, Stubbe J. 2011. Escherichia coli class Ib ribonucleotide reductase contains a dimanganese (III)

-tyrosyl radical cofactor in vivo. Biochemistry 50:1672–1681. DOI: https://doi.org/10.1021/bi101881d, PMID:

21250660

Edgar RC. 2004. Muscle: A multiple sequence alignment method with reduced time and space complexity. BMC

Bioinformatics 5:113. DOI: https://doi.org/10.1186/1471-2105-5-113, PMID: 15318951

Fehér T. 2008. Microbial gene essentiality: Protocols and bioinformatics. Humana.

Fraser CM, Casjens S, Huang WM, Sutton GG, Clayton R, Lathigra R, White O, Ketchum KA, Dodson R,

Hickey EK, Gwinn M, Dougherty B, Tomb JF, Fleischmann RD, Richardson D, Peterson J, Kerlavage AR,

Quackenbush J, Salzberg S, Hanson M, et al. 1997. Genomic sequence of a Lyme disease spirochaete, Borrelia

burgdorferi. Nature 390:580–586. DOI: https://doi.org/10.1038/37551, PMID: 9403685

Gilroy R, Ravi A, Getino M, Pursley I, Horton DL, Alikhan N-­F, Baker D, Gharbi K, Hall N, Watson M,

Adriaenssens EM, Foster-­Nyarko E, Jarju S, Secka A, Antonio M, Oren A, Chaudhuri RR, La Ragione R,

Hildebrand F, Pallen MJ. 2021. Extensive microbial diversity within the chicken gut microbiome revealed by

metagenomics and culture. PeerJ 9:e10941. DOI: https://doi.org/10.7717/peerj.10941, PMID: 33868800

Glass JI, Lefkowitz EJ, Glass JS, Heiner CR, Chen EY, Cassell GH. 2000. The complete sequence of the mucosal

pathogen Ureaplasma urealyticum. Nature 407:757–762. DOI: https://doi.org/10.1038/35037619, PMID:

11048724

Gupta RS, Bergström S. 2019. Distinction between borrelia and borreliella is more robustly supported by

molecular and phenotypic characteristics than all other neighbouring prokaryotic genera: response to margos’

et al. “the genus borrelia reloaded” (plos one 13(12): e0208432). PLOS ONE 14:e0221397. DOI: https://doi.​

org/10.1371/journal.pone.0221397, PMID: 31454394

Hershberg R, Petrov DA. 2010. Evidence that mutation is universally biased towards at in bacteria. PLOS

Genetics 6:e1001115. DOI: https://doi.org/10.1371/journal.pgen.1001115, PMID: 20838599

Hofer A, Crona M, Logan DT, Sjöberg B-­M. 2012. Dna building blocks: Keeping control of manufacture. Critical

Reviews in Biochemistry and Molecular Biology 47:50–63. DOI: https://doi.org/10.3109/10409238.2011.​

630372, PMID: 22050358

Horinouchi N, Ogawa J, Kawano T, Sakai T, Saito K, Matsumoto S, Sasaki M, Mikami Y, Shimizu S. 2006a.

Biochemical retrosynthesis of 2’-deoxyribonucleosides from glucose, acetaldehyde, and a nucleobase. Applied

Microbiology and Biotechnology 71:615–621. DOI: https://doi.org/10.1007/s00253-005-0205-5, PMID:

16283293

Arras et al. eLife 2023;12:e83845. DOI: https://doi.org/10.7554/eLife.83845

22 of 24

Evolutionary Biology | Microbiology and Infectious Disease

Research article

Horinouchi N, Ogawa J, Kawano T, Sakai T, Saito K, Matsumoto S, Sasaki M, Mikami Y, Shimizu S. 2006b.

One-­Pot microbial synthesis of 2’-deoxyribonucleoside from glucose, acetaldehyde, and a nucleobase.

Biotechnology Letters 28:877–881. DOI: https://doi.org/10.1007/s10529-006-9019-5, PMID: 16786272

Huang L, Zhang Y, Du X, An R, Liang X. 2022. Escherichia coli can eat DNA as an excellent nitrogen source to

grow quickly. Frontiers in Microbiology 13:894849. DOI: https://doi.org/10.3389/fmicb.2022.894849

Jeong H, Barbe V, Lee CH, Vallenet D, Yu DS, Choi S-­H, Couloux A, Lee S-­W, Yoon SH, Cattolico L, Hur C-­G,

Park H-­S, Ségurens B, Kim SC, Oh TK, Lenski RE, Studier FW, Daegelen P, Kim JF. 2009. Genome sequences of

Escherichia coli B strains REL606 and BL21(DE3). Journal of Molecular Biology 394:644–652. DOI: https://doi.​

org/10.1016/j.jmb.2009.09.052, PMID: 19786035

Karlström HO. 1970. Inability of Escherichia coli B to incorporate added deoxycytidine, deoxyandenosine, and

deoxyguanosine into DNA. European Journal of Biochemistry 17:68–71. DOI: https://doi.org/10.1111/j.1432-​

1033.1970.tb01135.x, PMID: 4922330

Langmead B, Salzberg SL. 2012. Fast gapped-­read alignment with bowtie 2. Nature Methods 9:357–359. DOI:

https://doi.org/10.1038/nmeth.1923, PMID: 22388286

Lee H, Popodi E, Tang H, Foster PL. 2012. Rate and molecular spectrum of spontaneous mutations in the

bacterium Escherichia coli as determined by whole-­genome sequencing. PNAS 109:E2774–E2783. DOI:

https://doi.org/10.1073/pnas.1210309109, PMID: 22991466

Lenski RE, Rose MR, Simpson SC, Tadler SC. 1991. Long-­Term experimental evolution in Escherichia coli. I.

adaptation and divergence during 2,000 generations. The American Naturalist 138:1315–1341. DOI: https://​

doi.org/10.1086/285289

Lundin D., Torrents E, Poole AM, Sjöberg BM. 2009. RNRdb, a curated database of the universal enzyme family

ribonucleotide reductase, reveals a high level of misannotation in sequences deposited to GenBank. BMC

Genomics 10:589. DOI: https://doi.org/10.1186/1471-2164-10-589, PMID: 19995434

Lundin D, Gribaldo S, Torrents E, Sjöberg B-­M, Poole AM. 2010. Ribonucleotide reduction-­horizontal transfer of

a required function spans all three domains. BMC Evolutionary Biology 10:383. DOI: https://doi.org/10.1186/​

1471-2148-10-383, PMID: 21143941

Lundin D, Berggren G, Logan DT, Sjöberg BM. 2015. The origin and evolution of ribonucleotide reduction. Life

5:604–636. DOI: https://doi.org/10.3390/life5010604, PMID: 25734234

Margos G, Marosevic D, Cutler S, Derdakova M, Diuk-­Wasser M, Emler S, Fish D, Gray J, Hunfeldt K-­P, Jaulhac B,

Kahl O, Kovalev S, Kraiczy P, Lane RS, Lienhard R, Lindgren PE, Ogden N, Ornstein K, Rupprecht T, Schwartz I,

et al. 2017. There is inadequate evidence to support the division of the genus Borrelia. International Journal of

Systematic and Evolutionary Microbiology 67:1081–1084. DOI: https://doi.org/10.1099/ijsem.0.001717

Margos G, Gofton A, Wibberg D, Dangel A, Marosevic D, Loh S-­M, Oskam C, Fingerle V. 2018. The genus

Borrelia reloaded. PLOS ONE 13:e0208432. DOI: https://doi.org/10.1371/journal.pone.0208432, PMID:

30586413

Margos G, Fingerle V, Oskam C, Stevenson B, Gofton A. 2020. Comment on: gupta, 2019, distinction between

borrelia and borreliella is more robustly supported by molecular and phenotypic characteristics than all other

neighbouring prokaryotic genera: response to margos’ et al. “the genus borrelia reloaded” (plos one 13(12):

e0208432). plos one 14(8):e0221397. Ticks and Tick-­Borne Diseases 11:101320. DOI: https://doi.org/10.1016/j.​

ttbdis.2019.101320, PMID: 31722850

Martin JE, Imlay JA. 2011. The alternative aerobic ribonucleotide reductase of Escherichia coli, NrdEF, is a

manganese-­dependent enzyme that enables cell replication during periods of iron starvation. Molecular

Microbiology 80:319–334. DOI: https://doi.org/10.1111/j.1365-2958.2011.07593.x, PMID: 21338418

Miller C, Thomsen LE, Gaggero C, Mosseri R, Ingmer H, Cohen SN. 2004. Sos response induction by ß-lactams

and bacterial defense against antibiotic lethality. Science 305:1629–1631. DOI: https://doi.org/10.1126/​

science.1101630

Miller SC, Porcella SF, Raffel SJ, Schwan TG, Barbour AG. 2013. Large linear plasmids of Borrelia species that

cause relapsing fever. Journal of Bacteriology 195:3629–3639. DOI: https://doi.org/10.1128/JB.00347-13,

PMID: 23749977

Monje-­Casas F, Jurado J, Prieto-­Alamo MJ, Holmgren A, Pueyo C. 2001. Expression analysis of THE nrdhief

operon from Escherichia coli conditions that trigger the transcript level in vivo. The Journal of Biological

Chemistry 276:18031–18037. DOI: https://doi.org/10.1074/jbc.M011728200, PMID: 11278973

Neidhardt FC, Bloch PL, Smith DF. 1974. Culture medium for enterobacteria. Journal of Bacteriology 119:736–

747. DOI: https://doi.org/10.1128/jb.119.3.736-747.1974, PMID: 4604283

Nguyen-­Vo TP, Ko S, Ryu H, Kim JR, Kim D, Park S. 2020. Systems evaluation reveals novel transporter yohjk

renders 3-­hydroxypropionate tolerance in Escherichia coli. Scientific Reports 10:19064. DOI: https://doi.org/10.​

1038/s41598-020-76120-3, PMID: 33149261

Nordlund P, Reichard P. 2006. Ribonucleotide reductases. Annual Review of Biochemistry 75:681–706. DOI:

https://doi.org/10.1146/annurev.biochem.75.103004.142443, PMID: 16756507

Ogawa J, Saito K, Sakai T, Horinouchi N, Kawano T, Matsumoto S, Sasaki M, Mikami Y, Shimizu S. 2003. Microbial

production of 2-­deoxyribose 5-­phosphate from acetaldehyde and triosephosphate for the synthesis of

2’-deoxyribonucleosides. Bioscience, Biotechnology, and Biochemistry 67:933–936. DOI: https://doi.org/10.​

1271/bbb.67.933, PMID: 12784646

Omer S, Harlow TJ, Gogarten JP. 2017. Does sequence conservation provide evidence for biological function?

Trends in Microbiology 25:11–18. DOI: https://doi.org/10.1016/j.tim.2016.09.010, PMID: 27773523

Panosian TD, Nannemann DP, Watkins GR, Phelan VV, McDonald WH, Wadzinski BE, Bachmann BO, Iverson TM.

2011. Bacillus cereus phosphopentomutase is an alkaline phosphatase family member that exhibits an altered

Arras et al. eLife 2023;12:e83845. DOI: https://doi.org/10.7554/eLife.83845

23 of 24

Evolutionary Biology | Microbiology and Infectious Disease

Research article

entry point into the catalytic cycle. The Journal of Biological Chemistry 286:8043–8054. DOI: https://doi.org/​

10.1074/jbc.M110.201350, PMID: 21193409

Patching SG, Baldwin SA, Baldwin AD, Young JD, Gallagher MP, Henderson PJF, Herbert RB. 2005. The

nucleoside transport proteins, nupc and nupg, from Escherichia coli: specific structural motifs necessary for the

binding of ligands. Organic & Biomolecular Chemistry 3:462. DOI: https://doi.org/10.1039/b414739a

Pérez-­Brocal V, Gil R, Ramos S, Lamelas A, Postigo M, Michelena JM, Silva FJ, Moya A, Latorre A. 2006. A small

microbial genome: the end of a long symbiotic relationship? Science 314:312–313. DOI: https://doi.org/10.​

1126/science.1130441, PMID: 17038625

Poole AM, Logan DT, Sjöberg BM. 2002. The evolution of the ribonucleotide reductases: much ado about

oxygen. Journal of Molecular Evolution 55:180–196. DOI: https://doi.org/10.1007/s00239-002-2315-3, PMID:

12107594

Poole AM, Horinouchi N, Catchpole RJ, Si D, Hibi M, Tanaka K, Ogawa J. 2014. The case for an early biological

origin of DNA. Journal of Molecular Evolution 79:204–212. DOI: https://doi.org/10.1007/s00239-014-9656-6,

PMID: 25425102

Posey JE, Gherardini FC. 2000. Lack of a role for iron in the Lyme disease pathogen. Science 288:1651–1653.

DOI: https://doi.org/10.1126/science.288.5471.1651, PMID: 10834845

Racker E. 1951. Enzymatic synthesis of deoxypentose phosphate. Nature 167:408–409. DOI: https://doi.org/10.​

1038/167408b0, PMID: 14826776

Racker E. 1952. Enzymatic synthesis and breakdown of desoxyribose phosphate. Journal of Biological Chemistry

196:347–365. DOI: https://doi.org/10.1016/S0021-9258(18)55739-2

Reichard P. 1993. From RNA to DNA, why so many ribonucleotide reductases? Science 260:1773–1777. DOI:

https://doi.org/10.1126/science.8511586, PMID: 8511586

Rodrigues JV, Shakhnovich EI. 2019. Adaptation to mutational inactivation of an essential gene converges to an

accessible suboptimal fitness peak. eLife 8:e50509. DOI: https://doi.org/10.7554/eLife.50509, PMID: 31573512

Rose HR, Ghosh MK, Maggiolo AO, Pollock CJ, Blaesi EJ, Hajj V, Wei Y, Rajakovich LJ, Chang WC, Han Y, Hajj M,

Krebs C, Silakov A, Pandelia ME, Bollinger JM, Boal AK. 2018. Structural basis for superoxide activation of

flavobacterium johnsoniae class i ribonucleotide reductase and for radical initiation by its dimanganese

cofactor. Biochemistry 57:2679–2693. DOI: https://doi.org/10.1021/acs.biochem.8b00247, PMID: 29609464

Schoch CL, Ciufo S, Domrachev M, Hotton CL, Kannan S, Khovanskaya R, Leipe D, Mcveigh R, O’Neill K,

Robbertse B, Sharma S, Soussov V, Sullivan JP, Sun L, Turner S, Karsch-­Mizrachi I. 2020. Ncbi taxonomy: a

comprehensive update on curation, resources and tools. Database 2020:baaa062. DOI: https://doi.org/10.​

1093/database/baaa062, PMID: 32761142

Silva F, Lourenço O, Pina-­Vaz C, Rodrigues AG, Queiroz JA, Domingues FC. 2010. The use of DRAQ5 to monitor

intracellular DNA in Escherichia coli by flow cytometry. Journal of Fluorescence 20:907–914. DOI: https://doi.​

org/10.1007/s10895-010-0636-y, PMID: 20352307

Sintchak MD, Arjara G, Kellogg BA, Stubbe J, Drennan CL. 2002. The crystal structure of class II ribonucleotide

reductase reveals how an allosterically regulated monomer mimics a dimer. Nature Structural Biology 9:293–

300. DOI: https://doi.org/10.1038/nsb774, PMID: 11875520

Srinivas V, Lebrette H, Lundin D, Kutin Y, Sahlin M, Lerche M, Eirich J, Branca RMM, Cox N, Sjöberg B-­M,

Högbom M. 2018. Metal-­Free ribonucleotide reduction powered by a dopa radical in Mycoplasma pathogens.

Nature 563:416–420. DOI: https://doi.org/10.1038/s41586-018-0653-6, PMID: 30429545

Stevenson K, McVey AF, Clark IBN, Swain PS, Pilizota T. 2016. General calibration of microbial growth in

microplate readers. Scientific Reports 6:38828. DOI: https://doi.org/10.1038/srep38828, PMID: 27958314

Tamas I, Klasson L, Canbäck B, Näslund AK, Eriksson AS, Wernegreen JJ, Sandström JP, Moran NA,

Andersson SGE. 2002. 50 million years of genomic stasis in endosymbiotic bacteria. Science 296:2376–2379.

DOI: https://doi.org/10.1126/science.1071278, PMID: 12089438

Tauer A, Benner SA. 1997. The B12-­dependent ribonucleotide reductase from the archaebacterium

Thermoplasma acidophila: an evolutionary solution to the ribonucleotide reductase conundrum. PNAS

94:53–58. DOI: https://doi.org/10.1073/pnas.94.1.53, PMID: 8990160

Torrents E, Aloy P, Gibert I, Rodríguez-­Trelles F. 2002. Ribonucleotide reductases: divergent evolution of an

ancient enzyme. Journal of Molecular Evolution 55:138–152. DOI: https://doi.org/10.1007/s00239-002-2311-7,

PMID: 12107591

Torrents E. 2014. Ribonucleotide reductases: essential enzymes for bacterial life. Frontiers in Cellular and

Infection Microbiology 4:52. DOI: https://doi.org/10.3389/fcimb.2014.00052, PMID: 24809024

Wang L, Westberg J, Bölske G, Eriksson S. 2001. Novel deoxynucleoside-­phosphorylating enzymes in

mycoplasmas: evidence for efficient utilization of deoxynucleosides. Molecular Microbiology 42:1065–1073.

DOI: https://doi.org/10.1046/j.1365-2958.2001.02700.x, PMID: 11737647

Yao Z, Kahne D, Kishony R. 2012. Distinct single-­cell morphological dynamics under beta-­lactam antibiotics.

Molecular Cell 48:705–712. DOI: https://doi.org/10.1016/j.molcel.2012.09.016, PMID: 23103254

Zhong J, Skouloubris S, Dai Q, Myllykallio H, Barbour AG. 2006. Function and evolution of plasmid-­borne genes

for pyrimidine biosynthesis in Borrelia spp. Journal of Bacteriology 188:909–918. DOI: https://doi.org/10.1128/​

JB.188.3.909-918.2006, PMID: 16428394

Arras et al. eLife 2023;12:e83845. DOI: https://doi.org/10.7554/eLife.83845

24 of 24

...