Stability and apoptotic activity of recombinant human cytochrome c

Alina Olteanu, Chetan N. Patel, Matthew M. Dedmon, Scott Kennedy, Michael W. Linhoff, Camille M. Minder, Patrick R. Potts, Mohanish Deshmukh, Gary J. Pielak

Research output: Contribution to journalArticle

43 Citations (Scopus)

Abstract

An efficient system for producing human cytochrome c variants is important to help us understand the roles of this protein in biological processes relevant to human diseases including apoptosis and oxidative stress. Here, we describe an Escherichia coli expression system for producing recombinant human cytochrome c. We also characterize the structure, stability, and function of the protein and show its utility for studying apoptosis. Yields of greater than 8mg of pure protein per liter culture were attained. Circular dichroism spectropolarimetry studies show that the secondary and tertiary structures of the human protein are nearly identical to those of the horse protein, but the human protein is more stable than other eukaryotic cytochromes c. Furthermore, recombinant human cytochrome c is capable of inducing caspase-3 activity in a cell-free caspase activation assay. We use data from this assay along with data from the literature to define the apaf-1 binding site on human cytochrome c.

Original languageEnglish (US)
Pages (from-to)733-740
Number of pages8
JournalBiochemical and Biophysical Research Communications
Volume312
Issue number3
DOIs
StatePublished - Dec 19 2003

Fingerprint

Cytochromes c
Human Activities
Proteins
Assays
Apoptosis
Oxidative stress
Biological Phenomena
Caspases
Protein Stability
Caspase 3
Escherichia coli
Circular Dichroism
Tertiary Protein Structure
Horses
Chemical activation
Binding Sites
Oxidative Stress

Keywords

  • Apoptosis
  • Circular dichroism
  • Cytochrome c
  • Protein stability

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Olteanu, A., Patel, C. N., Dedmon, M. M., Kennedy, S., Linhoff, M. W., Minder, C. M., ... Pielak, G. J. (2003). Stability and apoptotic activity of recombinant human cytochrome c. Biochemical and Biophysical Research Communications, 312(3), 733-740. https://doi.org/10.1016/j.bbrc.2003.10.182

Stability and apoptotic activity of recombinant human cytochrome c. / Olteanu, Alina; Patel, Chetan N.; Dedmon, Matthew M.; Kennedy, Scott; Linhoff, Michael W.; Minder, Camille M.; Potts, Patrick R.; Deshmukh, Mohanish; Pielak, Gary J.

In: Biochemical and Biophysical Research Communications, Vol. 312, No. 3, 19.12.2003, p. 733-740.

Research output: Contribution to journalArticle

Olteanu, A, Patel, CN, Dedmon, MM, Kennedy, S, Linhoff, MW, Minder, CM, Potts, PR, Deshmukh, M & Pielak, GJ 2003, 'Stability and apoptotic activity of recombinant human cytochrome c', Biochemical and Biophysical Research Communications, vol. 312, no. 3, pp. 733-740. https://doi.org/10.1016/j.bbrc.2003.10.182
Olteanu, Alina ; Patel, Chetan N. ; Dedmon, Matthew M. ; Kennedy, Scott ; Linhoff, Michael W. ; Minder, Camille M. ; Potts, Patrick R. ; Deshmukh, Mohanish ; Pielak, Gary J. / Stability and apoptotic activity of recombinant human cytochrome c. In: Biochemical and Biophysical Research Communications. 2003 ; Vol. 312, No. 3. pp. 733-740.
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