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Biotechnology and Applied Biochemistry (2002) 36, (213–218) (Printed in Great Britain)

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The calcium-binding protein of Entamoeba histolytica as a fusion partner for expression of peptides in Escherichia coli

Honey Reddi*¹, Alok Bhattacharya† and Vijay Kumar*²

*Virology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi-110 067, India and †School of Life Sciences, Jawaharlal Nehru University, New Mehrauli Road, New Delhi-110 067, India

Key words: affinity purification, enterokinase cleavage, fusion protein, soluble expression.

Abbreviations used: CaBP, calcium-binding protein; CBP, CaBP cloning vector; MCS, multiple cloning site; ERS, enterokinase recognition site; IPTG, isopropylthiogalactoside; Ni-NTA, Ni²⁺-nitrilotriacetate; MEP, multiple-epitope polypeptide.

¹Present address: B-620, ENH Research Institute, 2650 Ridge Avenue, Evanston Hospital, Evanston, IL 60201-1718, U.S.A.

²To whom correspondence should be addressed (e-mail vijay@icgeb.res.in).

We describe the construction of an Escherichia coli expression vector, CBP that allows the C-terminal fusion of heterologous proteins to the calcium-binding protein (CaBP) of the parasitic protozoan Entamoeba histolytica. The intrinsic nature of this protein to remain soluble on heat treatment has been exploited in its use as a novel fusion partner. The presence of a histidine tag and an enterokinase recognition site, aid in the affinity purification and proteolytic cleavage of the fusion protein. The efficacy of the vector was tested using the preS1 region of the envelope protein of the hepatitis B virus. The CaBP-preS1 fusion protein partitioned in the soluble fraction on heat treatment and this facilitated its rapid purification.

Received 28 March 2002/24 July 2002; accepted 2 August 2002

Portland Press Ltd © 2002