Isolation of the Rhizobium leguminosarum NodF nodulation protein: NodF carries a 4'-phosphopantetheine prosthetic group.
Rhizobium species produce a protein product of the nodF gene that has a limited but recognizable homology to the well-characterized acyl carrier protein (ACP) of Escherichia coli. NodF functions together with NodE in generating a host-specific response to the plant host in the interchange of signals leading to the effective nodulation of roots (H.P. Spaink, J. Weinman, M.A. Djordjevic, C.A. Wijffelman, R.J.H. Okker, and B. J.J. Lugtenberg, EMBO J. 8:2811-2818, 1989; B. Scheres, C. van de Wiel, A. Zalensky, B. Horvath, H. Spaink, H. van Eck, F. Zwartkruis, A.M. Wolters, T. Gloudemans, A. van Kammen, and T. Bisseling, Cell 60:281-294, 1990). The nodFE region of Rhizobium leguminosarum has been cloned into a multicopy plasmid and has been shown in R. leguminosarum to code for a flavonoid-inducible protein that is effectively labeled by radioactive beta-alanine added to the growth medium. After purification, the labeled protein migrates as a single band with an apparent molecular weight of 5,000 during sodium dodecyl sulfate-polyacrylamide gel electrophoresis, more rapidly than E. coli ACP. In contrast, in native gels the protein is resolved into two bands, both identified as NodF by analysis of the amino terminus and both migrating more slowly than E. coli ACP. Pulse-chase experiments with labeled beta-alanine suggested that the slower-moving band may be the precursor of the faster band. The NodF protein carries a 4'-phosphopantetheine as a prosthetic group. A NodF fusion protein under the control of the lac promoter is expressed in E. coli and is labeled with beta-alanine, indicating that it is recognized by the ACP synthase of E. coli. The ACP phosphodiesterase of E. coli, which catalyzes the release of phosphopantetheine from E. coli ACP, does not remove phosphopantetheine from NodF.
Geiger
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Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115.
Spaink
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Kennedy
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D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D002850:Chromatography, Gel; D002874:Chromosome Mapping; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D010204:Pantetheine / Q000031:analogs & derivatives / Q000032:analysis; D012231:Rhizobium / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid
D001426:Bacterial Proteins; C068605:nodF protein, Rhizobium; D010204:Pantetheine; C003129:4'-phosphopantetheine