Characterization of the Human Calmodulin-Like Protein Expressed in Escherichia Coli

Johannes A. Rhyner, Markus Roller, Isabelle Durussel-Gerber, Jos A. Cox, Emanuel E. Strehler

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Abstract

The protein-coding region of an intronless human calmodulin-like gene [Koller, M., Strehler, E.E. (1988) FEBS Lett. 239, 121-128] has been inserted into a pKK233-2 expression vector, and the 148-residue, Mx = 16 800 human protein was purified to apparent homogeneity by phenyl-Sepharose affinity chromatography from cultures of Escherichia coli JM105 transformed with the recombinant vector. Several milligrams of the purified protein were obtained from 1 L of bacterial culture. A number of properties of human CLP were compared to those of bacterially expressed human calmodulin (CaM) and of bovine brain CaM. CLP showed a characteristic Ca2+-dependent electrophoretic mobility shift on SDS-polyacrylamide gels, although the magnitude of this shift was smaller than that observed with CaM. CLP was able to activate the 3',5'-cyclic nucleotide phosphodiesterase to the same Kma, as normal CaM, albeit with a 7-fold higher Kact. In contrast, the erythrocyte plasma membrane Ca2+-ATPase could only be stimulated to 62% of its maximal CaM-dependent activity by CLP. CLP was found to contain four Ca2+-binding sites with a mean affinity constant of 105 M_1, a value about 10-fold lower than that for CaM under comparable conditions. The highly tissue-specifically-expressed CLP represents a novel human Ca2+-binding protein showing characteristics of a CaM isoform.

Original languageEnglish (US)
Pages (from-to)12826-12832
Number of pages7
JournalBiochemistry
Volume31
Issue number51
DOIs
StatePublished - Feb 1 1992

ASJC Scopus subject areas

  • Biochemistry

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    Rhyner, J. A., Roller, M., Durussel-Gerber, I., Cox, J. A., & Strehler, E. E. (1992). Characterization of the Human Calmodulin-Like Protein Expressed in Escherichia Coli. Biochemistry, 31(51), 12826-12832. https://doi.org/10.1021/bi00166a017