Applied Microbiology and Biotechnology

, Volume 103, Issue 6, pp 2809–2820 | Cite as

Expression and purification of an ArsM-elastin-like polypeptide fusion and its enzymatic properties

  • Changdong Ke
  • Hui Xiong
  • Chungui ZhaoEmail author
  • Zhigang Zhang
  • Xiaolan Zhao
  • Christopher RensingEmail author
  • Guangya Zhang
  • Suping YangEmail author
Environmental biotechnology


Enzymes could act as a useful tool for environmental bioremediation. Arsenic (As) biomethylation, which can convert highly toxic arsenite [As(III)] into low-toxic volatile trimethylarsine, is considered to be an effective strategy for As removal from contaminated environments. As(III) S-adenosylmethyltransferase (ArsM) is a key enzyme for As methylation; its properties and preparation are crucial for its wide application. Currently, ArsM is usually purified as a His-tag fusion protein restricting widespread use due to high costs. In this study, to greatly reduce the cost and simplify the ArsM preparation process, an Elastin-like polypeptide (ELP) tag was introduced to construct an engineered Escherichia coli (ArsM-ELP). Consequently, a cost-effective and simple non-chromatographic purification approach could be used for ArsM purification. The enzymatic properties of ArsM-ELP were systematically investigated. The results showed that the As methylation rate of purified ArsM-ELP (> 35.49%) was higher than that of E. coli (ArsM-ELP) (> 10.39%) when exposed to 25 μmol/L and 100 μmol/L As(III), respectively. The purified ArsM-ELP was obtained after three round inverse transition cycling treatment in 2.0 mol/L NaCl at 32 °C for 10 min with the yield reaching more than 9.6% of the total protein. The optimal reaction temperature, pH, and time of ArsM-ELP were 30 °C, 7.5 and 30 min, respectively. The enzyme activity was maintained at over 50% at 45 °C for 12 h. The enzyme specific activity was 438.8 ± 2.1 U/μmol. ArsM-ELP had high selectivity for As(III). 2-Mercaptoethanol could promote enzyme activity, whereas SDS, EDTA, Fe2+, and Cu2+ inhibited enzyme activity, and Mg2+, Zn2+, Ca2+, and K+ had no significant effects on it.


Arsenic As(III) S-adenosylmethionine (SAM) methyltransferase Elastin-like polypeptide tag Rhodopseudomonas palustris 



This study was funded by National Marine Public Industry Research (No. 201505026), by Natural Science Foundation of Fujian Province (No. 2018J01049), and by Subsidized Project for Cultivating Postgraduates Innovative Ability in Scientific Research of Huaqiao University.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any study with human participants or animals performed by any of the authors.


  1. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254. CrossRefGoogle Scholar
  2. Carlin A, Shi W, Dey S, Rosen BP (1995) The ars operon of Escherichia coli confers arsenical and antimonial resistance. J Bacteriol 177(4):981–986. CrossRefPubMedPubMedCentralGoogle Scholar
  3. Chen J, Qin J, Zhu YG, de Lorenzo V, Rosen BP (2013) Engineering the soil bacterium Pseudomonas putida for arsenic methylation. Appl Environ Microbiol 79(14):4493–4495. CrossRefPubMedPubMedCentralGoogle Scholar
  4. Chen J, Sun GX, Wang XX, de Lorenzo V, Rosen BP, Zhu YG (2014) Volatilization of arsenic from polluted soil by Pseudomonas putida engineered for expression of the arsM arsenic(III) S-adenosine methyltransferase gene. Environ Sci Technol 48(17):10337–10344.
  5. Dong H, Xu WZ, Pillai JK, Packianathan C, Rosen BP (2015) High-throughput screening-compatible assays of As(III) S-adenosylmethionine methyltransferase activity. Anal Biochem 480:67–73. CrossRefPubMedPubMedCentralGoogle Scholar
  6. Guo YQ, Xue XM, Yan Y, Zhu YG, Yang GD, Ye J (2016) Arsenic methylation by an arsenite S-adenosylmethionine methyltransferase from Spirulina platensis. J Environ Sci 49:162–168. CrossRefGoogle Scholar
  7. Huang KZ, Li JJ, Li W, Ge HH, Wang WY, Zhang GY (2011) De novo design, non-chromatographic purification and salt-effect of elastin-like polypeptides. Chin J Biotechnol 27(4):653–658Google Scholar
  8. Huang K, Chen C, Shen QR, Rosen BP, Zhao FJ (2015) Genetically engineering Bacillus subtilis with a heat-resistant arsenite methyltransferase for bioremediation of arsenic-contaminated organic waste. Appl Environ Microbiol 81(19):6718–6724. CrossRefPubMedPubMedCentralGoogle Scholar
  9. Huang K, Chen C, Zhang J, Tang Z, Shen QR, Rosen BP, Zhao FJ (2016) Efficient arsenic methylation and volatilization mediated by a novel bacterium from an arsenic-contaminated paddy soil. Environ Sci Technol 50(12):6389–6396. CrossRefPubMedPubMedCentralGoogle Scholar
  10. Huang K, Xu Y, Packianathan C, Gao F, Chen C, Zhang J, Shen QR, Rosen BP, Zhao FJ (2018) Arsenic methylation by a novel ArsM As(III) S-adenosylmethionine methyltransferase that requires only two conserved cysteine residues. Mol Microbiol 107(2):265–276. CrossRefPubMedGoogle Scholar
  11. Ke CD, Zhao CG, Rensing C, Yang SP, Zhang Y (2018) Characterization of recombinant E. coli expressing arsR from Rhodopseudomonas palustris CGA009 that displays highly selective arsenic adsorption. Appl Microbiol Biotechnol 102(14):6247–6255. CrossRefPubMedGoogle Scholar
  12. Kosobokova EN, Skrypnik KA, Kosorukov VS (2016) Overview of fusion tags for recombinant proteins. Biochem Mosc 81(3):187–200. CrossRefGoogle Scholar
  13. Kuramata M, Sakakibara F, Kataoka R, Abe T, Asano M, Baba K, Takagi K, Ishikawa S (2015) Arsenic biotransformation by Streptomyces sp isolated from rice rhizosphere. Environ Microbiol 17(6):1897–1909. CrossRefPubMedGoogle Scholar
  14. Li CC, Zhang GY (2014) The fusions of elastin-like polypeptides and xylanase self-assembled into insoluble active xylanase particles. J Biotechnol 177:60–66. CrossRefPubMedGoogle Scholar
  15. Lim DW, Trabbic-Carlson K, MacKay JA, Chilkoti A (2007) Improved non-chromatographic purification of a recombinant protein by cationic elastin-like polypeptides. Biomacromolecules 8(5):1417–1424. CrossRefPubMedPubMedCentralGoogle Scholar
  16. Liu S, Zhang F, Chen J, Sun GX (2011) Arsenic removal from contaminated soil via biovolatilization by genetically engineered bacteria under laboratory conditions. J Environ Sci 23(9):1544–1550. CrossRefGoogle Scholar
  17. Liu Z, Rensing C, Rosen BP (2014) Resistance pathways for metalloids and toxic metals. In: Culotta V, Scott RA (eds) Metals in cells. Wiley, Hoboken, pp 429–442Google Scholar
  18. Meng XY, Qin J, Wang LH, Duan GL, Sun GX, Wu HL, Chu CC, Ling HQ, Rosen BP, Zhu YG (2011) Arsenic biotransformation and volatilization in transgenic rice. New Phytol 191(1):49–56. CrossRefPubMedPubMedCentralGoogle Scholar
  19. Meyer DE, Chilkoti A (1999) Purification of recombinant proteins by fusion with thermally-responsive polypeptides. Nat Biotechnol 17(11):1112–1115CrossRefPubMedGoogle Scholar
  20. Oliveira LHB, Ferreira NS, Oliveira A, Nogueira ARA, Gonzalez MH (2017) Evaluation of distribution and bioaccumulation of arsenic by ICP-MS in Tilapia (Oreochromis niloticus) cultivated in different environments. J Braz Chem Soc 28(12):2455–2463Google Scholar
  21. Qin J, Rosen BP, Zhang Y, Wang GJ, Franke S, Rensing C (2006) Arsenic detoxification and evolution of trimethylarsine gas by a microbial arsenite S-adenosylmethionine methyltransferase. Proc Natl Acad Sci U S A 103(7):2075–2080. CrossRefPubMedPubMedCentralGoogle Scholar
  22. Qin J, Lehr CR, Yuan CG, Le XC, McDermott TR, Rosen BP (2009) Biotransformation of arsenic by a Yellowstone thermoacidophilic eukaryotic alga. Proc Natl Acad Sci U S A 106(13):5213–5217. CrossRefPubMedPubMedCentralGoogle Scholar
  23. Rao MA, Scelza R, Acevedo F, Diez MC, Gianfreda L (2014) Enzymes as useful tools for environmental purposes. Chemosphere 107:145–162. CrossRefPubMedGoogle Scholar
  24. Reguera J, Urry DW, Parker TM, McPherson DT, Rodriguez-Cabello JC (2007) Effect of NaCl on the exothermic and endothermic components of the inverse temperature transition of a model elastin-like polymer. Biomacromolecules 8(2):354–358. CrossRefPubMedGoogle Scholar
  25. Rensing C, Rosen BP (2009) Heavy metals cycles (arsenic, mercury, selenium, others). In: Schaechter M (ed) Encyclopedia of microbiology. Elsevier, Oxford, pp 205–219CrossRefGoogle Scholar
  26. Roberts S, Dzuricky M, Chilkoti A (2015) Elastin-like polypeptides as models of intrinsically disordered proteins. FEBS Lett 589(19):2477–2486. CrossRefPubMedPubMedCentralGoogle Scholar
  27. Sauge-Merle S, Cuine S, Carrier P, Lecomte-Pradines C, Luu DT, Peltier G (2003) Enhanced toxic metal accumulation in engineered bacterial cells expressing Arabidopsis thaliana phytochelatin synthase. Appl Environ Microbiol 69(1):490–494. CrossRefPubMedPubMedCentralGoogle Scholar
  28. Singh S, Mulchandani A, Chen W (2008) Highly selective and rapid arsenic removal by metabolically engineered Escherichia coli cells expressing fucus vesiculosus metallothionein. Appl Environ Microbiol 74(9):2924–2927. CrossRefPubMedPubMedCentralGoogle Scholar
  29. Singh JS, Abhilash PC, Singh HB, Singh RP, Singh DP (2011) Genetically engineered bacteria: an emerging tool for environmental remediation and future research perspectives. Gene 480(1–2):1–9. CrossRefPubMedGoogle Scholar
  30. Trabbic-Carlson K, Liu L, Kim B, Chilkoti A (2004) Expression and purification of recombinant proteins from Escherichia coli: comparison of an elastin-like polypeptide fusion with an oligohistidine fusion. Protein Sci 13(12):3274–3284. CrossRefPubMedPubMedCentralGoogle Scholar
  31. Verma S, Verma PK, Meher AK, Dwivedi S, Bansiwal AK, Pande V, Srivastava PK, Verma PC, Tripathi RD, Chakrabarty D (2016) A novel arsenic methyltransferase gene of Westerdykella aurantiaca isolated from arsenic contaminated soil: phylogenetic, physiological, and biochemical studies and its role in arsenic bioremediation. Metallomics 8(3):344–353. CrossRefPubMedGoogle Scholar
  32. Wang PP, Sun GX, Jia Y, Meharg AA, Zhu YG (2014a) A review on completing arsenic biogeochemical cycle: microbial volatilization of arsines in environment. J Environ Sci 26(2):371–381. CrossRefGoogle Scholar
  33. Wang PP, Sun GX, Zhu YG (2014b) Identification and characterization of arsenite methyltransferase from an archaeon, Methanosarcina acetivorans C2A. Environ Sci Technol 48(21):12706–12713. CrossRefPubMedGoogle Scholar
  34. Xue XM, Ye J, Raber G, Francesconi KA, Li G, Gao H, Yan Y, Rensing C, Zhu YG (2017) Arsenic methyltransferase is involved in arsenosugar biosynthesis by providing DMA. Environ Sci Technol 51(3):1224–1230. CrossRefPubMedGoogle Scholar
  35. Ye J, Rensing C, Rosen BP, Zhu YG (2012) Arsenic biomethylation by photosynthetic organisms. Trends Plant Sci 17(3):155–162. CrossRefPubMedPubMedCentralGoogle Scholar
  36. Ye J, Chang Y, Yan Y, Xiong J, Xue XM, Yuan DX, Sun GX, Zhu YG, Miao W (2014) Identification and characterization of the arsenite methyltransferase from a protozoan, Tetrahyrnena pyriformis. Aquat Toxicol 149:50–57. CrossRefPubMedGoogle Scholar
  37. Yin XX, Chen J, Qin J, Sun GX, Rosen BP, Zhu YG (2011) Biotransformation and volatilization of arsenic by three photosynthetic cyanobacteria. Plant Physiol 156(3):1631–1638. CrossRefPubMedPubMedCentralGoogle Scholar
  38. Yuan CG, Lu XF, Qin J, Rosen BP, Le XC (2008) Volatile arsenic species released from Escherichia coli expressing the AsIII S-adenosylmethionine methyltransferase gene. Environ Sci Technol 42(9):3201–3206. CrossRefPubMedPubMedCentralGoogle Scholar
  39. Zhang J, Cao TT, Tang Z, Shen QR, Rosen BP, Zhao FJ (2015) Arsenic methylation and volatilization by arsenite S-adenosylmethionine methyltransferase in Pseudomonas alcaligenes NBRC14159. Appl Environ Microbiol 81(8):2852–2860. CrossRefPubMedPubMedCentralGoogle Scholar
  40. Zhang J, Xu Y, Cao TT, Chen J, Rosen BP, Zhao FJ (2017) Arsenic methylation by a genetically engineered Rhizobium-legume symbiont. Plant Soil 416(1–2):259–269. CrossRefPubMedPubMedCentralGoogle Scholar
  41. Zhao CG, Zhang Y, Chan ZH, Chen SC, Yang SP (2015) Insights into arsenic multi-operons expression and resistance mechanisms in Rhodopseudomonas palustris CGA009. Front Microbiol 6:8. CrossRefGoogle Scholar
  42. Zhou YT, Niu LL, Liu K, Yin SS, Liu WP (2018) Arsenic in agricultural soils across China: distribution pattern, accumulation trend, influencing factors, and risk assessment. Sci Total Environ 616:156–163. CrossRefPubMedGoogle Scholar
  43. Zhu YG, Sun GX, Lei M, Teng M, Liu YX, Chen NC, Wang LH, Carey AM, Deacon C, Raab A, Meharg AA, Williams PN (2008) High percentage inorganic arsenic content of mining impacted and nonimpacted Chinese rice. Environ Sci Technol 42(13):5008–5013. CrossRefPubMedGoogle Scholar
  44. Zhu YG, Yoshinaga M, Zhao FJ, Rosen BP (2014) Earth abides arsenic biotransformations. Annu Rev Earth Planet Sci 42:443–467. CrossRefPubMedPubMedCentralGoogle Scholar
  45. Zhu YG, Xue XM, Kappler A, Rosen BP, Meharg AA (2017) Linking genes to microbial biogeochemical cycling: lessons from arsenic. Environ Sci Technol 51(13):7326–7339. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Bioengineering and BiotechnologyHuaqiao UniversityXiamenChina
  2. 2.Institute of Environmental Microbiology, College of Resources and EnvironmentFujian Agriculture and Forestry UniversityFuzhouChina
  3. 3.Key Laboratory of Urban Environment and Health, Institute of Urban EnvironmentChinese Academy of SciencesXiamenChina

Personalised recommendations