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Title: Production and preliminary characterization of a recombinant triheme cytochrome c7 from Geobacter sulfurreducens in Escherichia coli.

Abstract

Multiheme cytochromes c have been found in a number of sulfate- and metal ion-reducing bacteria. Geobacter sulfurreducens is one of a family of microorganisms that oxidize organic compounds, with Fe(III) oxide as the terminal electron acceptor. A triheme 9.6 kDa cytochrome c{sub 7} from G. sulfurreducens is a part of the metal ion reduction pathway. We cloned the gene for cytochrome c{sub 7} and expressed it in Escherichia coli together with the cytochrome c maturation gene cluster, ccmABCDEFGH, on a separate plasmid. We designed two constructs, with and without an N-terminal His-tag. The untagged version provided a good yield (up to 6 mg/l of aerobic culture) of the fully matured protein, with all three hemes attached, while the N-terminal His-tag appeared to be detrimental for proper heme incorporation. The recombinant protein (untagged) is properly folded, it has the same molecular weight and displays the same absorption spectra, both in reduced and in oxidized forms, as the protein isolated from G. sulfurreducens and it is capable of reducing metal ions in vitro. The shape parameters for the recombinant cytochrome c{sub 7} determined by small angle X-ray scattering are in good agreement with the ones calculated from a homologous cytochrome c{sub 7}more » of known structure.« less

Authors:
; ; ;
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States)
Sponsoring Org.:
USDOE Office of Science (SC)
OSTI Identifier:
949505
Report Number(s):
ANL/BIO/JA-41590
Journal ID: ISSN 0006-3002; BBACAQ; TRN: US201012%%293
DOE Contract Number:  
DE-AC02-06CH11357
Resource Type:
Journal Article
Journal Name:
Biochim. Biophys. Acta
Additional Journal Information:
Journal Volume: 1554; Journal Issue: 3 ; Jul. 1, 2002; Journal ID: ISSN 0006-3002
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; 99 GENERAL AND MISCELLANEOUS//MATHEMATICS, COMPUTING, AND INFORMATION SCIENCE; ABSORPTION SPECTRA; BINDING ENERGY; CYTOCHROMES; ESCHERICHIA COLI; IN VITRO; MOLECULAR WEIGHT; ORGANIC COMPOUNDS; PRODUCTION

Citation Formats

Londer, Y. Y., Pokkuluri, P. R., Tiede, D. M., and Schiffer, M. Production and preliminary characterization of a recombinant triheme cytochrome c7 from Geobacter sulfurreducens in Escherichia coli.. United States: N. p., 2002. Web. doi:10.1016/S0005-2728(02)00244-X.
Londer, Y. Y., Pokkuluri, P. R., Tiede, D. M., & Schiffer, M. Production and preliminary characterization of a recombinant triheme cytochrome c7 from Geobacter sulfurreducens in Escherichia coli.. United States. doi:10.1016/S0005-2728(02)00244-X.
Londer, Y. Y., Pokkuluri, P. R., Tiede, D. M., and Schiffer, M. Mon . "Production and preliminary characterization of a recombinant triheme cytochrome c7 from Geobacter sulfurreducens in Escherichia coli.". United States. doi:10.1016/S0005-2728(02)00244-X.
@article{osti_949505,
title = {Production and preliminary characterization of a recombinant triheme cytochrome c7 from Geobacter sulfurreducens in Escherichia coli.},
author = {Londer, Y. Y. and Pokkuluri, P. R. and Tiede, D. M. and Schiffer, M.},
abstractNote = {Multiheme cytochromes c have been found in a number of sulfate- and metal ion-reducing bacteria. Geobacter sulfurreducens is one of a family of microorganisms that oxidize organic compounds, with Fe(III) oxide as the terminal electron acceptor. A triheme 9.6 kDa cytochrome c{sub 7} from G. sulfurreducens is a part of the metal ion reduction pathway. We cloned the gene for cytochrome c{sub 7} and expressed it in Escherichia coli together with the cytochrome c maturation gene cluster, ccmABCDEFGH, on a separate plasmid. We designed two constructs, with and without an N-terminal His-tag. The untagged version provided a good yield (up to 6 mg/l of aerobic culture) of the fully matured protein, with all three hemes attached, while the N-terminal His-tag appeared to be detrimental for proper heme incorporation. The recombinant protein (untagged) is properly folded, it has the same molecular weight and displays the same absorption spectra, both in reduced and in oxidized forms, as the protein isolated from G. sulfurreducens and it is capable of reducing metal ions in vitro. The shape parameters for the recombinant cytochrome c{sub 7} determined by small angle X-ray scattering are in good agreement with the ones calculated from a homologous cytochrome c{sub 7} of known structure.},
doi = {10.1016/S0005-2728(02)00244-X},
journal = {Biochim. Biophys. Acta},
issn = {0006-3002},
number = 3 ; Jul. 1, 2002,
volume = 1554,
place = {United States},
year = {2002},
month = {7}
}