Identification and functional analysis of enzymes required for precorrin-2 dehydrogenation and metal ion insertion in the biosynthesis of sirohaem and cobalamin in Bacillus megaterium

Raux, Evelyne; Leech, Helen K.; Beck, Richard; Schubert, Heidi; Santander, Patricio J.; Roessner, Charles A.; Scott, A. Ian; Martens, Jan H.; Jahn, Dieter; Thermes, Claude; Rambach, Alain; Warren, Martin J.

doi:10.1042/bj20021443

Cited by 95 publications

(73 citation statements)

References 37 publications

(64 reference statements)

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“…In bacteria, SAM-dependent Urogen III methyltransferase is identified from eubacteria (Blanche et al 1989) and Archae (Blanche et al 1991). In some bacteria including Bacilus megaterium, the transformation of dihydrosirohydrochlorin (precorrin-2) into siroheme is catalyzed by two separate enzymes called SirC (dihydrosirohydrochlorin dehydro-genase) and SirB (sirohydrochlorin ferrochelatase) (Johansson and Hederstedt 1999;Raux et al 2003). However, in the plastids of higher plants, these activities seem to be separated into three distinct enzymes.…”

Section: Siroheme Branchmentioning

confidence: 99%

Tetrapyrrole Metabolism inArabidopsis thaliana

Tanaka

Kobayashi²,

Masuda

2011

The Arabidopsis Book

218

232

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This manuscript is dedicated to Prof. Mamoru Mimuro of Kyoto University who passed away in Februay 2011.Higher plants produce four classes of tetrapyrroles, namely, chlorophyll (Chl), heme, siroheme, and phytochromobilin. In plants, tetrapyrroles play essential roles in a wide range of biological activities including photosynthesis, respiration and the assimilation of nitrogen/sulfur. All four classes of tetrapyrroles are derived from a common biosynthetic pathway that resides in the plastid. In this article, we present an overview of tetrapyrrole metabolism in Arabidopsis and other higher plants, and we describe all identified enzymatic steps involved in this metabolism. We also summarize recent findings on Chl biosynthesis and Chl breakdown. Recent advances in this field, in particular those on the genetic and biochemical analyses of novel enzymes, prompted us to redraw the tetrapyrrole metabolic pathways. In addition, we also summarize our current understanding on the regulatory mechanisms governing tetrapyrrole metabolism. The interactions of tetrapyrrole biosynthesis and other cellular processes including the plastid-to-nucleus signal transduction are discussed.

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Section: Siroheme Branchmentioning

confidence: 99%

Tetrapyrrole Metabolism inArabidopsis thaliana

Tanaka

Kobayashi²,

Masuda

2011

The Arabidopsis Book

218

232

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“…Hence, structures of the Salmonella enterica CbiK (Se-CbiK) (6) and the Bacillus subtilis HemH (7), which are responsible for the insertion of Co 2þ and Fe 2þ into cobalamin and heme respectively, are bilobal enzymes consisting of two alpha/beta domains with a pseudo two-fold similarity, where the main catalytic groups are found in the C-terminal domain of the proteins. In contrast, the Bacillus megaterium CbiX L and SirB enzymes insert Co 2þ and Fe 2þ into SHC in the biosynthesis of cobalamin and siroheme, respectively, but in this case the active site residues are located in the N-terminal region of these proteins (2,8,9). Thus, these enzymes have evolved by a process of gene duplication and fusion followed by maintenance of the active site residues in either the N-or C-terminal region of the protein.…”

mentioning

confidence: 99%

Evolution in a family of chelatases facilitated by the introduction of active site asymmetry and protein oligomerization

Romão

Ladakis

Lobo

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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The class II chelatases associated with heme, siroheme, and cobalamin biosynthesis are structurally related enzymes that insert a specific metal ion (Fe 2þ or Co 2þ ) into the center of a modified tetrapyrrole (protoporphyrin or sirohydrochlorin). The structures of two related class II enzymes, CbiX S from Archaeoglobus fulgidus and CbiK from Salmonella enterica, that are responsible for the insertion of cobalt along the cobalamin biosynthesis pathway are presented in complex with their metallated product. A further structure of a CbiK from Desulfovibrio vulgaris Hildenborough reveals how cobalt is bound at the active site. The crystal structures show that the binding of sirohydrochlorin is distinctly different to porphyrin binding in the protoporphyrin ferrochelatases and provide a molecular overview of the mechanism of chelation. The structures also give insights into the evolution of chelatase form and function. Finally, the structure of a periplasmic form of Desulfovibrio vulgaris Hildenborough CbiK reveals a novel tetrameric arrangement of its subunits that are stabilized by the presence of a heme b cofactor. Whereas retaining colbaltochelatase activity, this protein has acquired a central cavity with the potential to chaperone or transport metals across the periplasmic space, thereby evolving a new use for an ancient protein subunit.enzyme mechanism | tetrapyrrole biosynthesis

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“…In the Archaea, CbiX is found as a comparatively small enzyme containing between 120 and 145 amino acids and is termed CbiX S1 (3), whereas in bacteria the enzyme is approximately twice the size containing around 320 amino acids, and this long version of the protein is termed CbiX L ( Fig. 2) (4). CbiX S displays similarity with both the N and C termini of CbiX L , indicating that CbiX L has probably arisen from a gene duplication and subsequent fusion of cbiX S (3).…”

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Characterization of the Cobaltochelatase CbiXL

Leech

Raux

McLean

et al. 2003

Journal of Biological Chemistry

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CbiX is a cobaltochelatase required for the biosynthesis of vitamin B 12 and is found in Archaea as a short form (CbiX S containing 120 -145 amino acids) and in some bacteria as a longer version (CbiX L containing 300 -350 amino acids). Purification of either recombinant Bacillus megaterium or Synechocystis CbiX L in Escherichia coli, which is facilitated by the presence of a naturally occurring histidine-rich region of the protein, results in the isolation of a dark brown protein solution. The UV/ visible spectrum of the protein is consistent with the presence of a redox group, and the lack of definition within the spectrum is suggestive of a 4Fe-4S center. The presence of an iron-sulfur center was confirmed by EPR analysis of the proteins, which produces a pseudoaxial spectrum with g values at 2.04, 1.94, and 1.90. The EPR spectrum was absent at 70 K, an observation that is diagnostic of a 4Fe-4S center. Redox potentiometry coupled with optical spectroscopy allowed the midpoint potential of the redox center to be determined for the CbiX L from both B. megaterium and Synechocystis. Sequence analysis of CbiX L proteins reveals only two conserved cysteine residues within the CbiX L proteins, which are part of an MXCXXC motif. Mutagenesis of the two cysteines leads to loss of both the EPR spectrum and UV/visible spectral features of the Fe-S center in the protein, clearly indicating that these residues are involved in ligating the cofactor to the apoprotein possibly in a butterfly arrangement. The potential physiological role of the iron-sulfur center is discussed.Modified tetrapyrroles such as heme, chlorophyll, siroheme, and vitamin B 12 (cobalamin) (1) are synthesized along a branched biosynthetic pathway, where each branch has a specific metal ion chelatase for the insertion of the particular metal ion found in the center of the tetrapyrrole-derived macrocycle. CbiX is a cobaltochelatase that inserts cobalt into sirohydrochlorin during the biosynthesis of cobalamin (vitamin B 12 ) ( Fig. 1) (2). In the Archaea, CbiX is found as a comparatively small enzyme containing between 120 and 145 amino acids and is termed CbiX S1 (3), whereas in bacteria the enzyme is approximately twice the size containing around 320 amino acids, and this long version of the protein is termed CbiX L (Fig. 2) (4). CbiX S displays similarity with both the N and C termini of CbiX L , indicating that CbiX L has probably arisen from a gene duplication and subsequent fusion of cbiX S (3). CbiX L was first described in Bacillus megaterium and was shown to consist of 306 amino acids with a very unusual histidine-rich region toward the C terminus of the protein (2). More recently, a similar protein named SirB, found within a small siroheme biosynthetic operon, has been described that is capable of inserting ferrous iron into sirohydrochlorin during the biosynthesis of siroheme (4). SirB contains 266 amino acids and shares 40% identity with CbiX L . The difference in length between SirB and CbiX L is largely due to the presence of a C-terminal ...

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Identification and functional analysis of enzymes required for precorrin-2 dehydrogenation and metal ion insertion in the biosynthesis of sirohaem and cobalamin in Bacillus megaterium

Cited by 95 publications

References 37 publications

Tetrapyrrole Metabolism inArabidopsis thaliana

Tetrapyrrole Metabolism inArabidopsis thaliana

Evolution in a family of chelatases facilitated by the introduction of active site asymmetry and protein oligomerization

Characterization of the Cobaltochelatase CbiXL

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