Identification of triphosphoribosyl-dephospho-CoA as precursor of the citrate lyase prosthetic group.
The gamma-subunit of citrate lyase (EC 4.1.3.6) contains the prosthetic group 2'-(5"-phosphoribosyl)-3'-dephospho-CoA and serves as an acyl carrier protein (ACP). We recently showed that in Escherichia coli the proteins CitG and CitX are essential for holo-ACP synthesis and provided evidence that CitG catalyzes the formation of a prosthetic group precursor from ATP and dephospho-CoA, which is subsequently attached via phosphodiester linkage to apo-ACP by CitX. Here we prove that CitG indeed catalyzes the conversion of ATP and dephospho-CoA to adenine and 2'-(5"-triphosphoribosyl)-3'-dephospho-CoA, the predicted precursor of the prosthetic group. Furthermore, this precursor was transferred by CitX to apo-ACP, yielding holo-ACP. Thus, our proposed mechanism for holo-ACP synthesis could be verified.
Schneider
K
K
Institut für Mikrobiologie, Eidgenössische Technische Hochschule Zürich, Switzerland.
Dimroth
P
P
Bott
M
M
D001275:ATP Citrate (pro-S)-Lyase / Q000737:chemistry / Q000378:metabolism*; D000213:Acyl Carrier Protein / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001051:Apoenzymes / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003065:Coenzyme A / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004792:Enzyme Precursors / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D020035:Holoenzymes / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000213:Acyl Carrier Protein; D001051:Apoenzymes; D004792:Enzyme Precursors; D020035:Holoenzymes; C015787:dephosphocoenzyme A; D000255:Adenosine Triphosphate; D001275:ATP Citrate (pro-S)-Lyase; D003065:Coenzyme A