Post-translational modification in the archaea: structural characterization of multi-enzyme complex lipoylation

Mareike G. Posner, Abhishek Upadhyay, Susan Crennell, Andrew J. A. Watson, Stephen Dorus, Michael J. Danson, Stefan Bagby

Research output: Contribution to journalArticle

8 Citations (Scopus)
72 Downloads (Pure)

Abstract

Lipoylation, the covalent attachment of lipoic acid to 2-oxoacid dehydrogenase multi-enzyme complexes, is essential for metabolism in aerobic bacteria and eukarya. In Escherichia coli, lipoylation is catalysed by lipoate protein ligase (LplA) or by lipoic acid synthetase (LipA) and lipoyl(octanoyl) transferase (LipB) combined. Whereas bacterial and eukaryotic LplAs comprise a single, two-domain protein, archaeal LplA function typically involves two proteins, LplA-N and LplA-C. In the thermophilic archaeon Thermoplasma acidophilum, LplA-N and LplA-C are encoded by overlapping genes in inverted orientation (lpla-c is upstream of lpla-n). The structure of Thermoplasma acidophilum LplA-N is known, but the structure of LplA-C and its role in lipoylation are unknown. We have determined the structures of the substrate-free LplA-N+LplA-C complex and the dihydrolipoyl acyltransferase lipoyl domain (E2lipD) that is lipoylated by LplA-N+LplA-C, and carried out biochemical analyses of this archaeal lipoylation system. Our data
reveal the following: LplA-C is disordered but folds upon association with LplA-N; LplA-C induces a conformational change in LplA-N involving substantial shortening of a loop that could repress catalytic activity of isolated LplA-N; the adenylate binding region of LplA-N+LplA-C includes two helices rather than the purely loop structure of varying order observed in other LplA structures; LplA-N+LplA-C and E2lipD do not interact in the absence of substrate; LplA-N+LplA-C undergoes a conformational change (the details of which are currently undetermined) during lipoylation; LplA-N+LplA-C can utilize octanoic acid as well as lipoic acid as substrate. The elucidated functional inter-dependence of LplA-N and LplA-C is consistent with their evolutionary co-retention in archaeal genomes.
Original languageEnglish
Pages (from-to)415-425
Number of pages11
JournalBiochemical Journal
Volume449
Issue number2
Early online date1 Nov 2012
DOIs
Publication statusPublished - 15 Jan 2013

Fingerprint

Lipoylation
Archaea
Post Translational Protein Processing
Enzymes
Protein C
lipoate-protein ligase
Multienzyme Complexes
Thioctic Acid
Thermoplasma

Keywords

  • binding-induced folding
  • lipoate protein ligase
  • lipoyl domain
  • NMR spectroscopy
  • protein-protein interaction
  • X-ray crystallography

Cite this

Post-translational modification in the archaea: structural characterization of multi-enzyme complex lipoylation. / Posner, Mareike G.; Upadhyay, Abhishek; Crennell, Susan; Watson, Andrew J. A.; Dorus, Stephen; Danson, Michael J.; Bagby, Stefan.

In: Biochemical Journal, Vol. 449, No. 2, 15.01.2013, p. 415-425.

Research output: Contribution to journalArticle

Posner, Mareike G. ; Upadhyay, Abhishek ; Crennell, Susan ; Watson, Andrew J. A. ; Dorus, Stephen ; Danson, Michael J. ; Bagby, Stefan. / Post-translational modification in the archaea: structural characterization of multi-enzyme complex lipoylation. In: Biochemical Journal. 2013 ; Vol. 449, No. 2. pp. 415-425.
@article{64fb4982274e43c2b9826ff76ac13574,
title = "Post-translational modification in the archaea: structural characterization of multi-enzyme complex lipoylation",
abstract = "Lipoylation, the covalent attachment of lipoic acid to 2-oxoacid dehydrogenase multi-enzyme complexes, is essential for metabolism in aerobic bacteria and eukarya. In Escherichia coli, lipoylation is catalysed by lipoate protein ligase (LplA) or by lipoic acid synthetase (LipA) and lipoyl(octanoyl) transferase (LipB) combined. Whereas bacterial and eukaryotic LplAs comprise a single, two-domain protein, archaeal LplA function typically involves two proteins, LplA-N and LplA-C. In the thermophilic archaeon Thermoplasma acidophilum, LplA-N and LplA-C are encoded by overlapping genes in inverted orientation (lpla-c is upstream of lpla-n). The structure of Thermoplasma acidophilum LplA-N is known, but the structure of LplA-C and its role in lipoylation are unknown. We have determined the structures of the substrate-free LplA-N+LplA-C complex and the dihydrolipoyl acyltransferase lipoyl domain (E2lipD) that is lipoylated by LplA-N+LplA-C, and carried out biochemical analyses of this archaeal lipoylation system. Our data reveal the following: LplA-C is disordered but folds upon association with LplA-N; LplA-C induces a conformational change in LplA-N involving substantial shortening of a loop that could repress catalytic activity of isolated LplA-N; the adenylate binding region of LplA-N+LplA-C includes two helices rather than the purely loop structure of varying order observed in other LplA structures; LplA-N+LplA-C and E2lipD do not interact in the absence of substrate; LplA-N+LplA-C undergoes a conformational change (the details of which are currently undetermined) during lipoylation; LplA-N+LplA-C can utilize octanoic acid as well as lipoic acid as substrate. The elucidated functional inter-dependence of LplA-N and LplA-C is consistent with their evolutionary co-retention in archaeal genomes.",
keywords = "binding-induced folding, lipoate protein ligase, lipoyl domain, NMR spectroscopy, protein-protein interaction, X-ray crystallography",
author = "Posner, {Mareike G.} and Abhishek Upadhyay and Susan Crennell and Watson, {Andrew J. A.} and Stephen Dorus and Danson, {Michael J.} and Stefan Bagby",
year = "2013",
month = "1",
day = "15",
doi = "10.1042/BJ20121150",
language = "English",
volume = "449",
pages = "415--425",
journal = "Biochemical Journal",
issn = "0264-6021",
publisher = "Portland Press Ltd.",
number = "2",

}

TY - JOUR

T1 - Post-translational modification in the archaea: structural characterization of multi-enzyme complex lipoylation

AU - Posner, Mareike G.

AU - Upadhyay, Abhishek

AU - Crennell, Susan

AU - Watson, Andrew J. A.

AU - Dorus, Stephen

AU - Danson, Michael J.

AU - Bagby, Stefan

PY - 2013/1/15

Y1 - 2013/1/15

N2 - Lipoylation, the covalent attachment of lipoic acid to 2-oxoacid dehydrogenase multi-enzyme complexes, is essential for metabolism in aerobic bacteria and eukarya. In Escherichia coli, lipoylation is catalysed by lipoate protein ligase (LplA) or by lipoic acid synthetase (LipA) and lipoyl(octanoyl) transferase (LipB) combined. Whereas bacterial and eukaryotic LplAs comprise a single, two-domain protein, archaeal LplA function typically involves two proteins, LplA-N and LplA-C. In the thermophilic archaeon Thermoplasma acidophilum, LplA-N and LplA-C are encoded by overlapping genes in inverted orientation (lpla-c is upstream of lpla-n). The structure of Thermoplasma acidophilum LplA-N is known, but the structure of LplA-C and its role in lipoylation are unknown. We have determined the structures of the substrate-free LplA-N+LplA-C complex and the dihydrolipoyl acyltransferase lipoyl domain (E2lipD) that is lipoylated by LplA-N+LplA-C, and carried out biochemical analyses of this archaeal lipoylation system. Our data reveal the following: LplA-C is disordered but folds upon association with LplA-N; LplA-C induces a conformational change in LplA-N involving substantial shortening of a loop that could repress catalytic activity of isolated LplA-N; the adenylate binding region of LplA-N+LplA-C includes two helices rather than the purely loop structure of varying order observed in other LplA structures; LplA-N+LplA-C and E2lipD do not interact in the absence of substrate; LplA-N+LplA-C undergoes a conformational change (the details of which are currently undetermined) during lipoylation; LplA-N+LplA-C can utilize octanoic acid as well as lipoic acid as substrate. The elucidated functional inter-dependence of LplA-N and LplA-C is consistent with their evolutionary co-retention in archaeal genomes.

AB - Lipoylation, the covalent attachment of lipoic acid to 2-oxoacid dehydrogenase multi-enzyme complexes, is essential for metabolism in aerobic bacteria and eukarya. In Escherichia coli, lipoylation is catalysed by lipoate protein ligase (LplA) or by lipoic acid synthetase (LipA) and lipoyl(octanoyl) transferase (LipB) combined. Whereas bacterial and eukaryotic LplAs comprise a single, two-domain protein, archaeal LplA function typically involves two proteins, LplA-N and LplA-C. In the thermophilic archaeon Thermoplasma acidophilum, LplA-N and LplA-C are encoded by overlapping genes in inverted orientation (lpla-c is upstream of lpla-n). The structure of Thermoplasma acidophilum LplA-N is known, but the structure of LplA-C and its role in lipoylation are unknown. We have determined the structures of the substrate-free LplA-N+LplA-C complex and the dihydrolipoyl acyltransferase lipoyl domain (E2lipD) that is lipoylated by LplA-N+LplA-C, and carried out biochemical analyses of this archaeal lipoylation system. Our data reveal the following: LplA-C is disordered but folds upon association with LplA-N; LplA-C induces a conformational change in LplA-N involving substantial shortening of a loop that could repress catalytic activity of isolated LplA-N; the adenylate binding region of LplA-N+LplA-C includes two helices rather than the purely loop structure of varying order observed in other LplA structures; LplA-N+LplA-C and E2lipD do not interact in the absence of substrate; LplA-N+LplA-C undergoes a conformational change (the details of which are currently undetermined) during lipoylation; LplA-N+LplA-C can utilize octanoic acid as well as lipoic acid as substrate. The elucidated functional inter-dependence of LplA-N and LplA-C is consistent with their evolutionary co-retention in archaeal genomes.

KW - binding-induced folding

KW - lipoate protein ligase

KW - lipoyl domain

KW - NMR spectroscopy

KW - protein-protein interaction

KW - X-ray crystallography

UR - http://www.scopus.com/inward/record.url?scp=84871436925&partnerID=8YFLogxK

UR - http://dx.doi.org/10.1042/BJ20121150

U2 - 10.1042/BJ20121150

DO - 10.1042/BJ20121150

M3 - Article

VL - 449

SP - 415

EP - 425

JO - Biochemical Journal

JF - Biochemical Journal

SN - 0264-6021

IS - 2

ER -