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@ARTICLE{Schneider:26938,
author = {Schneider, K. and Dimroth, B. M. and Bott, M.},
title = {{B}iosynthesis of the prosthetic group of citrate lyase},
journal = {Biochemistry},
volume = {39},
issn = {0006-2960},
address = {Columbus, Ohio},
publisher = {American Chemical Society},
reportid = {PreJuSER-26938},
pages = {9438 - 9450},
year = {2000},
note = {Record converted from VDB: 12.11.2012},
abstract = {Citrate lyase (EC 4.1.3.6) catalyzes the cleavage of
citrate to acetate and oxaloacetate and is composed of three
subunits (alpha, beta, and gamma). The gamma-subunit serves
as an acyl carrier protein (ACP) and contains the prosthetic
group 2'-(5' '-phosphoribosyl)-3'-dephospho-CoA, which is
attached via a phosphodiester linkage to serine-14 in the
enzyme from Klebsiella pneumoniae. In this work, we
demonstrate by genetic and biochemical studies with citrate
lyase of Escherichia coli and K. pneumoniae that the
conversion of apo-ACP into holo-ACP is dependent on the two
proteins, CitX (20 kDa) and CitG (33 kDa). In the absence of
CitX, only apo-ACP was synthesized in vivo, whereas in the
absence of CitG, an adenylylated ACP was produced, with the
AMP residue attached to serine-14. The adenylyltransferase
activity of CitX could be verified in vitro with purified
CitX and apo-ACP plus ATP as substrates. Besides ATP, CTP,
GTP, and UTP also served as nucleotidyl donors in vitro,
showing that CitX functions as a nucleotidyltransferase. The
conversion of apo-ACP into holo-ACP was achieved in vitro by
incubation of apo-ACP with CitX, CitG, ATP, and
dephospho-CoA. ATP could not be substituted with GTP, CTP,
UTP, ADP, or AMP. In the absence of CitG or dephospho-CoA,
AMP-ACP was formed. Remarkably, it was not possible to
further convert AMP-ACP to holo-ACP by subsequent incubation
with CitG and dephospho-CoA. This demonstrates that AMP-ACP
is not an intermediate during the conversion of apo- into
holo-ACP, but results from a side activity of CitX that
becomes effective in the absence of its natural substrate.
Our results indicate that holo-ACP formation proceeds as
follows. First, a prosthetic group precursor [presumably
2'-(5' '-triphosphoribosyl)-3'-dephospho-CoA] is formed from
ATP and dephospho-CoA in a reaction catalyzed by CitG.
Second, holo-ACP is formed from apo-ACP and the prosthetic
group precursor in a reaction catalyzed by CitX.},
keywords = {Acyl Carrier Protein: biosynthesis / Amino Acid Sequence /
Apoproteins: biosynthesis / Carbon-Sulfur Ligases:
biosynthesis / Carbon-Sulfur Ligases: genetics / Coenzyme A:
biosynthesis / Enzyme Precursors: metabolism / Escherichia
coli: enzymology / Escherichia coli: genetics / Genes,
Bacterial / Molecular Sequence Data / Multienzyme Complexes:
biosynthesis / Multienzyme Complexes: chemistry /
Multienzyme Complexes: genetics / Multigene Family / Operon
/ Oxo-Acid-Lyases: biosynthesis / Oxo-Acid-Lyases: chemistry
/ Oxo-Acid-Lyases: genetics / Structure-Activity
Relationship / Acyl Carrier Protein (NLM Chemicals) /
Apoproteins (NLM Chemicals) / Enzyme Precursors (NLM
Chemicals) / Multienzyme Complexes (NLM Chemicals) /
dephosphocoenzyme A (NLM Chemicals) / Coenzyme A (NLM
Chemicals) / Oxo-Acid-Lyases (NLM Chemicals) / citrate
(pro-3S)-lyase (NLM Chemicals) / Carbon-Sulfur Ligases (NLM
Chemicals) / long-chain-fatty-acid-(acyl-carrier-protein)
ligase (NLM Chemicals) / citrate (pro-3S)-lyase ligase (NLM
Chemicals) / J (WoSType)},
cin = {IBT},
ddc = {570},
cid = {I:(DE-Juel1)VDB184},
pnm = {Entwicklung von Mikroorganismen für die Herstellung von
Primärmetaboliten},
pid = {G:(DE-Juel1)FUEK91},
shelfmark = {Biochemistry $\&$ Molecular Biology},
typ = {PUB:(DE-HGF)16},
pubmed = {pmid:10924139},
UT = {WOS:000088593300042},
url = {https://juser.fz-juelich.de/record/26938},
}