Cloning and characterization of mitochondrial 5-formyltetrahydrofolate cycloligase from higher plants

Sanja Roje; Machhindra T Janave; Michael J Ziemak; Andrew D Hanson

doi:10.1074/jbc.M205632200

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Cloning and characterization of mitochondrial 5-formyltetrahydrofolate cycloligase from higher plants

Journal article

Open access

Peer reviewed

Cloning and characterization of mitochondrial 5-formyltetrahydrofolate cycloligase from higher plants

Sanja Roje, Machhindra T Janave, Michael J Ziemak and Andrew D Hanson

The Journal of biological chemistry, Vol.277(45), pp.42748-42754

11/08/2002

DOI: https://doi.org/10.1074/jbc.M205632200

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https://hdl.handle.net/2376/103878

PMID: 12207015

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Abstract

Recombinant Proteins - metabolism

Amino Acid Sequence

Mitochondria - enzymology

Arabidopsis - enzymology

Escherichia coli - enzymology

Molecular Weight

Carbon-Nitrogen Ligases - metabolism

Humans

DNA, Complementary - genetics

Molecular Sequence Data

Substrate Specificity

Phylogeny

DNA Primers

Carbon-Nitrogen Ligases - genetics

Sequence Homology, Amino Acid

Sequence Alignment

Animals

Escherichia coli - genetics

Base Sequence

Cloning, Molecular

Lycopersicon esculentum - genetics

5-Formyltetrahydrofolate cycloligase (5-FCL) catalyzes the conversion of 5-formyltetrahydrofolate (5-CHO-H(4)PteGlu(n)) to 5,10-methenyltetrahydrofolate and is considered to be the main means whereby 5-CHO-H(4)PteGlu(n) is metabolized in mammals, yeast, and bacteria. 5-CHO-H(4)PteGlu(n) is known to occur in plants and to be highly abundant in leaf mitochondria. Genomics-based approaches identified Arabidopsis and tomato cDNAs encoding proteins homologous to 5-FCLs of other organisms but containing N-terminal extensions with the features of mitochondrial targeting peptides. These homologs were shown to have 5-FCL activity by characterizing recombinant enzymes produced in Escherichia coli and by functional complementation of a yeast fau1 mutation with the Arabidopsis 5-FCL cDNA. The recombinant Arabidopsis enzyme is active as a monomer, prefers the penta- to the monoglutamyl form of 5-CHO-H(4)PteGlu(n), and has kinetic properties broadly similar to those of 5-FCLs from other organisms. Enzyme assays and immunoblot analyses indicated that 5-FCL is located predominantly if not exclusively in plant mitochondria and that the mature, active enzyme lacks the putative targeting sequence. Serine hydroxymethyltransferase (SHMT) from plant mitochondria was shown to be inhibited by 5-CHO-H(4)PteGlu(n) as are SHMTs from other organisms. Since mitochondrial SHMT is crucial to photorespiration, 5-FCL may help prevent 5-CHO-H(4)PteGlu(n) from reaching levels that would inhibit this process. Consistent with this possibility, 5-FCL activity was far higher in leaf mitochondria than root mitochondria.

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Title: Cloning and characterization of mitochondrial 5-formyltetrahydrofolate cycloligase from higher plants
Creators: Sanja Roje - Horticultural Sciences Department, University of Florida, Gainesville, Florida 32611, USA
Machhindra T Janave
Michael J Ziemak
Andrew D Hanson
Publication Details: The Journal of biological chemistry, Vol.277(45), pp.42748-42754
Academic Unit: Biological Chemistry, Institute of
Publisher: United States
Identifiers: 99900546725901842
Language: English
Resource Type: Journal article