Please use this identifier to cite or link to this item: https://doi.org/10.1002/jctb.280590111
Title: Purification and renaturation of recombinant human lymphotoxin (tumour necrosis factor beta) expressed in Escherichia coli as inclusion bodies
Authors: Jin, H.
Uddin, M.S. 
Huang, Y.L. 
Teo, W.K. 
Keywords: E coli
Inclusion bodies
Purification
Refolding
rhTNF-β
Issue Date: Jan-1994
Citation: Jin, H., Uddin, M.S., Huang, Y.L., Teo, W.K. (1994-01). Purification and renaturation of recombinant human lymphotoxin (tumour necrosis factor beta) expressed in Escherichia coli as inclusion bodies. Journal of Chemical Technology and Biotechnology 59 (1) : 67-72. ScholarBank@NUS Repository. https://doi.org/10.1002/jctb.280590111
Abstract: High level expression of recombinant human tumour necrosis factor (rhTNF-β) in Escherichia coli results in the formation of two portions of protein, namely soluble active protein and insoluble protein which is inactive and aggregates in the form of inclusion bodies (IBs). In this study, a procedure for purification and renaturation of rhTNF-β from inclusion bodies has been designed and verified experimentally with a product purity of more than 90% and a recovery of about 30%. The procedure includes washing of IBs with specific wash buffer (Triton X-100/EDTA/lysozyme/PMSF), their solubilization with 8 mol dm-3 alkaline urea, purification with ion-exchange columns, refolding with renaturation buffer and finally concentration and desalination with an ultrafiltration membrane. The characteristics of the renatured protein were identical with those of purified protein from the soluble fraction as demonstrated by (1) SDS-PAGE, (2) cytotoxic activity on mouse L929 cells, (3) N-terminal amino acid sequence, and (4) gel filtration chromatography.
Source Title: Journal of Chemical Technology and Biotechnology
URI: http://scholarbank.nus.edu.sg/handle/10635/91665
ISSN: 02682575
DOI: 10.1002/jctb.280590111
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