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AtJ1, a mitochondrial homologue of theEscherichia coli DnaJ protein

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Abstract

The nucleotide sequence of a cDNA clone fromArabidopsis thaliana ecotype Columbia was determined, and the corresponding amino sequence deduced. The open reading frame encodes a protein, AtJ1, of 368 residues with a molecular mass of 41 471 Da and an isoelectric point of 9.2. The predicted sequence contains regions homologous to the J- and cysteine-rich domains ofEscherichia coli DnaJ, but the glycine/phenylalanine-rich region is not present. Based upon Southern analysis,Arabidopsis appears to have a singleatJ1 structural gene. A single species of mRNA, of 1.5 kb, was detected whenArabidopsis poly(A)+ RNA was hybridized with theatJ1 cDNA. The function ofatJ1 was tested by complementation of adnaJ deletion mutant ofE. coli, allowing growth in minimal medium at 44°C. The AtJ1 protein was expressed inE. coli as a fusion with the maltose binding protein. This fusion protein was purified by amylose affinity chromatography, then cleaved by digestion with the activated factor X protease. The recombinant AtJ1 protein was purified to electrophoretic homogeneity.In vitro, recombinant AtJ1 stimulated the ATPase activity of bothE. coli DnaK and maize endosperm cytoplasmic Stress70. The deduced amino acid sequence of AtJ1 contains a potential mitochondrial targeting sequence at the N-terminus. Radioactive recombinant AtJ1 was synthesized inE. coli and purified. When the labeled protein was incubated with intact pea cotyledon mitochondria, it was imported and proteolytically processed in a reaction that depended upon an energized mitochondrial membrane.

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Abbreviations

MBP:

maltose binding protein

PCR:

polymerase chain reaction

Stress70c:

the cytosolic member of the 70 kDA family of stress-related proteins

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Author notes

  1. Barbara Kroczynska

    Present address: Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawinskiego 5A, 02-106, Warszawa, Poland

  2. Rengang Zhou

    Present address: Institute of Agro-Physics, Plant Physiology and Biochemistry, Hebei Academy of Agricultural Sciences, Shijiazhuang, People's Republic of China

Authors and Affiliations

  1. Mycotoxin Research Unit, USDA, Agricultural Research Service, National Center for Agricultural Utilization Research, 61604-3902, Peoria, IL, USA

    Barbara Kroczynska, Rengang Zhou & Jan A. Miernyk

  2. Department of Biological and Nutritional Sciences, the University, NE1 7RU, Newcastle upon Tyne, UK

    Clifford Wood

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  1. Barbara Kroczynska
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  3. Clifford Wood
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  4. Jan A. Miernyk
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Kroczynska, B., Zhou, R., Wood, C. et al. AtJ1, a mitochondrial homologue of theEscherichia coli DnaJ protein. Plant Mol Biol 31, 619–629 (1996). https://doi.org/10.1007/BF00042234

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  • DOI: https://doi.org/10.1007/BF00042234

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