A 113-amino acid fragment of CD4 produced in Escherichia coli blocks human immunodeficiency virus-induced cell fusion

B. H. Chao, D. S. Costopoulos, T. Curiel, J. M. Bertonis, P. Chisholm, C. Williams, R. T. Schooley, J. J. Rosa, R. A. Fisher, J. M. Maraganore

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33 Scopus citations


A gene encoding a 113-amino acid, NH2-terminal fragment of CD4, rsT4,113, was constructed and expressed in Escherichia coli under the control of the tryptophan operon promoter. Following induction, rsT4.113 is produced at 5-10% of total E. coli protein, and it is found in inclusion bodies. The protein is purified in two steps under denaturing and reducing conditions. Solubilized rsT4.113 is first purified on a column of Q-Sepharose to remove low molecular weight contaminants and then purified to > 95% homogeneity by gel filtration. Renaturation of rsT4.113 is achieved at approximately 20% yield by dilution and dialysis. High performance liquid chromatography analysis of renatured rsT4.113 reveals a < 15% contaminant of reduced protein. Purified and renatured rsT4.113 contains epitopes for both OKT4a and Leu3a, anti-CD4 monoclonal antibodies which block CD4-gp120 association, but lacks measurable affinity toward a nonblocking anti-CD4 monoclonal antibody, OKT4. By comparison to a longer form (375 amino acids) of recombinant soluble T4 produced in mammalian cells that contains the entire extracellular domain, rsT4.113 has a comparable affinity for binding to OKT4a and Leu3a in a radioimmunoassay. Analysis of antiviral activity of rsT4.113 demonstrates that the E. coli-derived protein inhibits human immunodeficiency virus-induced syncytium formation with an IC50 of 5-10 μg/ml. These data demonstrate that the human immunodeficiency virus-binding domain of CD4 is localized within the NH2-terminal 113 amino acids of CD4 and is contained within a structure homologous to the κ variable-like domain of immunoglobulins.

Original languageEnglish (US)
Pages (from-to)5812-5817
Number of pages6
JournalJournal of Biological Chemistry
Issue number10
StatePublished - 1989
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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