Cloning, purification and enzymatic characterization of recombinant human superoxide dismutase 1 (hSOD1) expressed in Escherichia coli

• Authors: Feng Lin , Dandan Yan , Yawen Chen , Fletcher E , Haifeng Shi , Bangxing Han , Yang Zhou
• Languages of publication: EN

Abstracts

EN

Superoxide dismutase 1 (SOD1) is a metalloenzyme that catalyzes the disproportionation of superoxide into molecular oxygen and hydrogen peroxide. In this study, the human SOD1 (hSOD1) gene was cloned, expressed and purified. The hSOD1 gene was amplified from a pool of Bxpc3 cell cDNAs by PCR and cloned into expression vector pET-28a (+). The recombinant soluble hSOD1 was expressed in E. coli BL21 (DE3) at 37°C and purified using nickel column affinity chromatography. Soluble hSOD1 was produced with a yield of 5.9 μg/mL medium. As metal ions can have a certain influence on protein structure and activity, we researched the influences of different concentrations of Cu2+ and Zn2+ on hSOD1 activity at induction and the time of activity detection. The results implied that Cu2+ and Zn2+ do not enhance SOD1 expression and solubility; they can, however, improve the catalytic activity at induction. Meanwhile, Cu2+ and Zn2+ also enhanced the enzyme activity at the time of detection. Furthermore, most other bivalent cations had the potential to replace Zn2+ and Cu2+, and also improved enzyme activity at the time of detection.

Keywords

EN

superoxide dismutase 1; Escherichia coli; soluble expression; metal ions; catalytic activity

• Publisher: Polish Biochemical Society
• Journal: Acta Biochimica Polonica
• Year: 2018
• Volume: 65
• Issue: 2
• Pages: 235-240

Dates

published

2018

received

2017-11-08

revised

2018-02-15

accepted

2018-02-17

(unknown)

2018-05-30

Contributors

author

Feng Lin

• School of Food and Biological Engineering, Jiangsu University, Zhenjiang 212013, P. R. China
• Institute of Life Sciences, Jiangsu University, Zhenjiang 212013, P. R. China
• Key Laboratory of Healthy Freshwater Aquaculture, Ministry of Agriculture, Zhejiang Institute of Freshwater Fisheries, Huzhou 313001, P. R. China

author

Dandan Yan

• Institute of Life Sciences, Jiangsu University, Zhenjiang 212013, P. R. China

author

Yawen Chen

• Institute of Life Sciences, Jiangsu University, Zhenjiang 212013, P. R. China

author

Fletcher E

• Institute of Life Sciences, Jiangsu University, Zhenjiang 212013, P. R. China

author

Haifeng Shi

• Institute of Life Sciences, Jiangsu University, Zhenjiang 212013, P. R. China

author

Bangxing Han

• College of Biological and Pharmaceutical Engineering, West Anhui University, Moon Island, Lu'an 237012, P. R. China

author

Yang Zhou

• Institute of Life Sciences, Jiangsu University, Zhenjiang 212013, P. R. China

References

• Ahl IM, Lindberg MJ, Tibell LA (2004) Coexpression of yeast copper chaperone (yCCS) and CuZn-superoxide dismutases in Escherichia coli yields protein with high copper contents. Protein Expr Purif 37: 311-319.doi: 10.1016/j.pep.2004.06.006.
• Banci L, Barbieri L, Bertini I, Cantini F, Luchinat E (2011) In-cell NMR in E. coli to monitor maturation steps of hSOD1. PLoS One 6: e23561.doi: 10.1371/journal.pone.0023561.
• Crow JP, Sampson JB, Zhuang Y, Thompson JA, Beckman JS (1997) Decreased zinc affinity of amyotrophic lateral sclerosis-associated superoxide dismutase mutants leads to enhanced catalysis of tyrosine nitration by peroxynitrite. J Neurochem 69: 1936-1944.
• Eiamphungporn W, Yainoy S, Prachayasittikul V (2016) Enhancement of solubility and specific activity of a Cu/Zn superoxide dismutase by Co-expression with a copper chaperone in Escherichia coli. Iran J Biotechnol 14: 243-249.doi: 10.15171/ijb.1465.
• Fujii J, Myint T, Seo HG, Kayanoki Y, Ikeda Y, Taniguchi N (1995) Characterization of wild-type and amyotrophic lateral sclerosis-related mutant CuZn-superoxide dismutases overproduced in baculovirus-infected insect cells. J Neurochem 64: 1456-1461.
• Geraghty P, Baumlin N, Salathe MA, Foronjy RF, D'Armiento JM (2016) Glutathione Peroxidase-1 suppresses the unfolded protein response upon cigarette smoke exposure. Mediators Inflamm 2016: 9461289.doi: 10.1155/2016/9461289.
• Ghneim HK (2016) The kinetics of the effect of manganese supplementation on SOD2 activity in senescent human fibroblasts. Eur Rev Med Pharmacol Sci 20: 1866-1880.
• Girotto S, Cendron L, Bisaglia M, Tessari I, Mammi S, Zanotti G, Bubacco L (2014) DJ-1 is a copper chaperone acting on SOD1 activation. J Biol Chem 289: 10887-10899.doi: 10.1074/jbc.M113.535112.
• Haddad NI, Yuan Q (2005) Purification and some properties of CuZn superoxide dismutase from Radix lethospermi seed kind of Chinese traditional medicine. J Chromatogr B Analyt Technol Biomed Life Sci 818: 123-131.doi: 10.1016/j.jchromb.2004.12.010.
• Hartman JR, Geller T, Yavin Z, Bartfeld D, Kanner D, Aviv H, Gorecki M (1986) High-level expression of enzymatically active human Cu/Zn superoxide dismutase in Escherichia coli. Proc Natl Acad Sci U S A 83: 7142-7146.
• Hayward LJ, Rodriguez JA, Kim JW, Tiwari A, Goto JJ, Cabelli DE, Valentine JS, Brown RH Jr (2002) Decreased metallation and activity in subsets of mutant superoxide dismutases associated with familial amyotrophic lateral sclerosis. J Biol Chem 277: 15923-15931.doi: 10.1074/jbc.M112087200.
• Hole PS, Darley RL, Tonks A (2011) Do reactive oxygen species play a role in myeloid leukemias? Blood 117: 5816-5826.doi: 10.1182/blood-2011-01-326025.
• Huo J, Shi H, Yao Q, Chen H, Wang L, Chen K (2010) Cloning and purification of recombinant silkworm dihydrolipoamide dehydrogenase expressed in Escherichia coli. Protein Expr Purif 72: 95-100.doi: 10.1016/j.pep.2010.01.014.
• Johnson P (2002) Antioxidant enzyme expression in health and disease: effects of exercise and hypertension. Comp Biochem Physiol C Toxicol Pharmacol 133: 493-505.
• Kilic N, Taslipinar YM, Guney Y, Tekin E, Onuk E (2014) An investigation into the serum thioredoxin superoxide dismutase malondialdehyde and advanced oxidation protein products in patients with breast cancer. Ann Surg Oncol 21: 4139-4143.doi: 10.1245/s10434-014-3859-3.
• Leitch JM, Jensen LT, Bouldin SD, Outten CE, Hart PJ, Culotta VC (2009) Activation of Cu,Zn-superoxide dismutase in the absence of oxygen and the copper chaperone CCS. J Biol Chem 284: 21863-21871.doi: 10.1074/jbc.M109.000489.
• Li HT, Jiao M, Chen J, Liang Y (2010) Roles of zinc and copper in modulating the oxidative refolding of bovine copper zinc superoxide dismutase. Acta Biochim Biophys Sin (Shanghai) 42: 183-194.
• Lin C, Zeng H, Lu J, Xie Z, Sun W, Luo C, Ding J, Yuan S, Geng M, Huang M (2015) Acetylation at lysine 71 inactivates superoxide dismutase 1 and sensitizes cancer cells to genotoxic agents. Oncotarget 6: 20578-20591.doi: 10.18632/oncotarget.3987.
• Ming LJ, Valentine JS (2014) Insights into SOD1-linked amyotrophic lateral sclerosis from NMR studies of Ni(2+)- and other metal-ion-substituted wild-type copper-zinc superoxide dismutases. J Biol Inorg Chem 19: 647-657.doi: 10.1007/s00775-014-1126-5.
• Nordlund A, Leinartaite L, Saraboji K, Aisenbrey C, Grobner G, Zetterstrom P, Danielsson J, Logan DT, Oliveberg M (2009) Functional features cause misfolding of the ALS-provoking enzyme SOD1. Proc Natl Acad Sci U S A 106: 9667-9672.doi: 10.1073/pnas.0812046106.
• Park DH, Yoon S-YH Nam HG, Park JM (2002) Expression of functional human-cytosolic Cu/Zn superoxide dismutase in transgenic tobacco. Biotechnology Letters 24: 681-686
• Rumfeldt JA, Lepock JR, Meiering EM (2009) Unfolding and folding kinetics of amyotrophic lateral sclerosis-associated mutant CuZn superoxide dismutases. J Mol Biol 385: 278-298.doi: 10.1016/j.jmb.2008.10.003.
• Sangwan S, Zhao A, Adams KL, Jayson CK, Sawaya MR, Guenther EL, Pan AC, Ngo J, Moore DM, Soriaga AB (2017) Atomic structure of a toxic oligomeric segment of SOD1 linked to amyotrophic lateral sclerosis (ALS). Proc Natl Acad Sci USA 114: 8770-8775.
• Shaw BF, Valentine JS (2007) How do ALS-associated mutations in superoxide dismutase 1 promote aggregation of the protein? Trends Biochem Sci 32: 78-85.doi: 10.1016/j.tibs.2006.12.005.
• Shih LY, Liou TH, Chao JC, Kau HN, Wu YJ, Shieh MJ, Yeh CY, Han BC (2006) Leptin, superoxide dismutase, and weight loss: initial leptin predicts weight loss. Obesity (Silver Spring) 14: 2184-2192.doi: 10.1038/oby.2006.256.
• Swalley SE, Fulghum JR, Chambers SP (2006) Screening factors effecting a response in soluble protein expression: formalized approach using design of experiments. Anal Biochem 351: 122-127.doi: 10.1016/j.ab.2005.11.046.
• Vasina JA, Baneyx F (1997) Expression of aggregation-prone recombinant proteins at low temperatures: a comparative study of the Escherichia coli cspA and tac promoter systems. Protein Expr Purif 9: 211-218.doi: 10.1006/prep.1996.0678.
• Vats P, Sagar N, Singh TP, Banerjee M (2015) Association of superoxide dismutases (SOD1 and SOD2) and glutathione peroxidase 1 (GPx1) gene polymorphisms with type 2 diabetes mellitus. Free Radic Res 49: 17-24.doi: 10.3109/10715762.2014.971782.
• Wittung-Stafshede P (2004) Role of cofactors in folding of the blue-copper protein azurin. Inorg Chem 43: 7926-7933.doi: 10.1021/ic049398g.
• Wu CY, Steffen J, Eide DJ (2009) Cytosolic superoxide dismutase (SOD1) is critical for tolerating the oxidative stress of zinc deficiency in yeast. PLoS One 4: e7061.doi: 10.1371/journal.pone.0007061.
• Yoo HY, Kim SS, Rho HM (1999) Overexpression and simple purification of human superoxide dismutase (SOD1) in yeast and its resistance to oxidative stress. J Biotechnol 68: 29-35.
• Zhang K, Zhang Y, Zi J, Xue X, Wan Y (2017) Production of human Cu,Zn SOD with higher activity and lower toxicity in E. coli via mutation of free cysteine residues. Biomed Res Int 2017: 4817376.doi: 10.1155/2017/4817376.
• Zhou Y, Yuan S, Liu Q, Yan D, Wang Y, Gao L, Han J, Shi H (2017) Synchronized purification and immobilization of his-tagged beta-glucosidase via Fe3O4/PMG core/shell magnetic nanoparticles. Sci Rep 7: 41741.doi: 10.1038/srep41741.

Identifiers

DOI

10.18388/abp.2017_2350