1H NMR study of the interaction of ATP with Escherichia coli RNA polymerase containing in vivo-incorporated Co(II).
The DNA-dependent RNA polymerase containing two intrinsic cobalt ions (Co2-RPase) instead of the naturally occurring zinc was purified from Escherichia coli cells grown in zinc-depleted, cobalt-enriched media. Longitudinal NMR relaxation rates of the H2 and H8 protons of ATP were measured in the absence and presence of up to 92 microM Co2-RPase. No enhancement of the proton relaxation rates was observed in the presence of cobalt-containing enzyme, suggesting that the ATP substrate does not undergo rapid exchange at a site close to either of the intrinsic cobalt ions. This result is in contrast to that previously observed when Co2+ was incorporated into RPase by an in vitro procedure involving partial urea denaturation of the protein.
Panth
H
H
Department of Pharmacological Sciences, State University of New York, Stony Brook 11794.
Brenner
M C
MC
Wu
F Y
FY
D000255:Adenosine Triphosphate / Q000737:chemistry*; D002413:Cations, Divalent; D003035:Cobalt / Q000737:chemistry*; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000302:isolation & purification; D004926:Escherichia coli / Q000201:enzymology*; D009682:Magnetic Resonance Spectroscopy; D013053:Spectrophotometry; D015032:Zinc
D002413:Cations, Divalent; D003035:Cobalt; D000255:Adenosine Triphosphate; D012321:DNA-Directed RNA Polymerases; D015032:Zinc