Calmodulin as a versatile tag for antibody fragments

Dario Neri, Claudia De Lalla, Heike Petrul, Paolo Neri, Greg Winter

Research output: Contribution to journalArticle

49 Citations (Scopus)

Abstract

Calmodulin is a highly acidic protein (net charge -24 at pH 8.0 in the absence of calcium) that binds to peptide and organic ligands with high affinity (Ka>109 M-1) in a calcium-dependent manner. We have exploited these properties to develop calmodulin as a versatile tag for antibody fragments. Fusions of calmodulin with single chain Fv fragments (scFv) could be expressed by secretion from bacteria in good yield (5-15 mg/l in shaker flasks), and purified from periplasmic lysates or broth to homogeneity in a single step, either by binding to anion-exchange resin (DEAE-Sephadex), or to an organic ligand of calmodulin (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide-agarose). The antibody fusions could be detected by binding of fluorescently labeled peptide ligands, as illustrated by their use in confocal microscopy, fluorescent activated cell sorting and "band shift" gel electrophoresis. Moreover, the interaction between calmodulin and peptide ligands could provide a means of heterodimerization of proteins, as illustrated by the assembly of an antibody-calmodulin fusion with maltose binding protein tagged with a peptide ligand of calmodulin.

Original languageEnglish
Pages (from-to)373-377
Number of pages5
JournalNature Biotechnology
Volume13
Issue number4
Publication statusPublished - 1995

Fingerprint

Calmodulin
Immunoglobulin Fragments
Antibodies
Ligands
Peptides
Fusion reactions
Calcium
Anion Exchange Resins
Maltose-Binding Proteins
Immunoglobulin Variable Region
Proteins
Maltose
DEAE-Dextran
Single-Chain Antibodies
Confocal microscopy
Electrophoresis
Sorting
Confocal Microscopy
Sepharose
Ion exchange

ASJC Scopus subject areas

  • Microbiology
  • Biotechnology

Cite this

Neri, D., De Lalla, C., Petrul, H., Neri, P., & Winter, G. (1995). Calmodulin as a versatile tag for antibody fragments. Nature Biotechnology, 13(4), 373-377.

Calmodulin as a versatile tag for antibody fragments. / Neri, Dario; De Lalla, Claudia; Petrul, Heike; Neri, Paolo; Winter, Greg.

In: Nature Biotechnology, Vol. 13, No. 4, 1995, p. 373-377.

Research output: Contribution to journalArticle

Neri, D, De Lalla, C, Petrul, H, Neri, P & Winter, G 1995, 'Calmodulin as a versatile tag for antibody fragments', Nature Biotechnology, vol. 13, no. 4, pp. 373-377.
Neri D, De Lalla C, Petrul H, Neri P, Winter G. Calmodulin as a versatile tag for antibody fragments. Nature Biotechnology. 1995;13(4):373-377.
Neri, Dario ; De Lalla, Claudia ; Petrul, Heike ; Neri, Paolo ; Winter, Greg. / Calmodulin as a versatile tag for antibody fragments. In: Nature Biotechnology. 1995 ; Vol. 13, No. 4. pp. 373-377.
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