Two-step purification of Cyanophora ferredoxin and its identification in soluble protein preparations by isoelectric focusing.
Cyanophora paradoxa ferredoxin is encoded by (cyano-)plastidic DNA, in contrast to those of all other photosynthetic eukaryotes investigated so far. In the present study we report (i) the rapid purification of a chloroplast-type [2Fe-2S] ferredoxin in a two-step procedure by DEAE-Sephadex and Mono Q ion-exchange chromatography; (ii) the biochemical characterization of the purified ferredoxin by electrophoretic separation methods on a microscale; and (iii) a qualitative and quantitative ferredoxin detection method in the femtomole range that allows densitometry, semidry immunoblotting, identification of ferredoxin in soluble cell protein preparations, and analysis of protein biosynthesis from cyanoplast poly(A)- RNA in vivo and in vitro. These fast micromethods should be useful for screening phototrophic species containing ferredoxins encoded by nonnuclear DNA.
Bayer
M G
MG
Institut für Allgemeine Botanik und Pfanzenphysiologie, Tübingen, Germany.
Gebhart
U B
UB
Maier
T L
TL
Schenk
H E
HE
D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004269:DNA, Bacterial / Q000235:genetics; D056890:Eukaryota / Q000737:chemistry*; D005288:Ferredoxins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D007525:Isoelectric Focusing; D007526:Isoelectric Point; D015388:Organelles / Q000737:chemistry; D014176:Protein Biosynthesis; D012995:Solubility; D013053:Spectrophotometry
D004269:DNA, Bacterial; D005288:Ferredoxins