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Oxaloacetate decarboxylase of Archaeoglobus fulgidus: cloning of genes and expression in Escherichia coli

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Abstract

Archaeoglobus fulgidus harbors three consecutive and one distantly located gene with similarity to the oxaloacetate decarboxylase Na+ pump of Klebsiella pneumoniae (KpOadGAB). The water-soluble carboxyltransferase (AfOadA) and the biotin protein (AfOadC) were readily synthesized in Escherichia coli, but the membrane-bound subunits AfOadB and AfOadG were not. AfOadA was affinity purified from inclusion bodies after refolding and AfOadC was affinity purified from the cytosol. Isolated AfOadA catalyzed the carboxyltransfer from [4-14C]-oxaloacetate to the prosthetic biotin group of AfOadC or the corresponding biotin domain of KpOadA. Conversely, the carboxyltransferase domain of KpOadA exhibited catalytic activity not only with its pertinent biotin domain but also with AfOadC.

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Abbreviations

Af:

Archaeoglobus fulgidus

AHT:

Anhydrotetracycline

DTT:

Dithiothreitol

IPTG:

Isopropyl-β-d-thiogalactopyranoside

Kp:

Klebsiella pneumoniae

LDAO:

N,N-dimethyldodecylamine-N-oxide

MBP:

Maltose binding protein

OAD:

Oxaloacetate decarboxylase

OadA:

Oxaloacetate decarboxylase α-subunit

OadB:

Oxaloacetate decarboxylase β-subunit

OadC:

Oxaloacetate decarboxylase δ-subunit

OadG:

Oxaloacetate decarboxylase γ-subunit

PPS:

3-(1-pyridino)-1-propanesulfate

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Acknowledgments

This work was supported by Swiss National Science Foundation. We thank Prof. Dr M. Thomm and Dr Harald Huber (Archaeenzentrum Universität Regensburg) for their generous gift of large amounts of A. fulgidus cells.

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Authors and Affiliations

  1. Institut für Mikrobiologie der ETH Zürich, ETH-Zentrum, Schmelzbergstrasse 7, 8092, Zürich, Switzerland

    Pius Dahinden, Klaas M. Pos, Malgorzata Taralczak & Peter Dimroth

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  1. Pius Dahinden
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  2. Klaas M. Pos
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  3. Malgorzata Taralczak
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  4. Peter Dimroth
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Correspondence to Peter Dimroth.

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Dahinden, P., Pos, K.M., Taralczak, M. et al. Oxaloacetate decarboxylase of Archaeoglobus fulgidus: cloning of genes and expression in Escherichia coli. Arch Microbiol 182, 414–420 (2004). https://doi.org/10.1007/s00203-004-0706-0

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  • DOI: https://doi.org/10.1007/s00203-004-0706-0

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