An FMN hydrolase is fused to a riboflavin kinase homolog in plants

Francisco J Sandoval; Sanja Roje

doi:10.1074/jbc.M500350200

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An FMN hydrolase is fused to a riboflavin kinase homolog in plants

Journal article

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Peer reviewed

An FMN hydrolase is fused to a riboflavin kinase homolog in plants

Francisco J Sandoval and Sanja Roje

The Journal of biological chemistry, Vol.280(46), pp.38337-38345

11/18/2005

DOI: https://doi.org/10.1074/jbc.M500350200

Handle:

https://hdl.handle.net/2376/107727

PMID: 16183635

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Abstract

Temperature

Arabidopsis - enzymology

Molecular Weight

Phosphotransferases (Alcohol Group Acceptor) - chemistry

Humans

Molecular Sequence Data

Chromatography, High Pressure Liquid

Phylogeny

Time Factors

Plant Proteins - chemistry

Cloning, Molecular

Escherichia coli - metabolism

Gene Expression Regulation, Plant

Catalysis

Expressed Sequence Tags

Hydrolases - metabolism

Protein Structure, Tertiary

Amino Acid Sequence

Computational Biology

Recombinant Proteins - chemistry

Flavin Mononucleotide - chemistry

Reverse Transcriptase Polymerase Chain Reaction

Chromatography

Phosphotransferases (Alcohol Group Acceptor) - metabolism

Arabidopsis - genetics

Sequence Homology, Amino Acid

Magnesium - chemistry

Animals

Edetic Acid - chemistry

Protein Binding

Cytosol - metabolism

Hydrolases - chemistry

Kinetics

DNA, Complementary - metabolism

Hydrogen-Ion Concentration

Riboflavin kinases catalyze synthesis of FMN from riboflavin and ATP. These enzymes have to date been cloned from bacteria, yeast, and mammals, but not from plants. Bioinformatic approaches suggested that diverse plant species, including many angiosperms, two gymnosperms, a moss (Physcomitrella patens), and a unicellular green alga (Chlamydomonas reinhardtii), encode proteins that are homologous to riboflavin kinases of yeast and mammals, but contain an N-terminal domain that belongs to the haloacid dehalogenase superfamily of enzymes. The Arabidopsis homolog of these proteins was cloned by RT-PCR, and was shown to have riboflavin kinase and FMN hydrolase activities by characterizing the recombinant enzyme produced in Escherichia coli. Both activities of the purified recombinant Arabidopsis enzyme (AtFMN/FHy) increased when the enzyme assays contained 0.02% Tween 20. The FMN hydrolase activity of AtFMN/FHy greatly decreased when EDTA replaced Mg(2+) in the assays, as expected for a member of the Mg(2+)-dependent haloacid dehalogenase family. The functional overexpression of the individual domains in E. coli establishes that the riboflavin kinase and FMN hydrolase activities reside, respectively, in the C-terminal (AtFMN) and N-terminal (AtFHy) domains of AtFMN/FHy. Biochemical characterization of AtFMN/FHy, AtFMN, and AtFHy shows that the riboflavin kinase and FMN hydrolase domains of AtFMN/FHy can be physically separated, with little change in their kinetic properties.

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Title: An FMN hydrolase is fused to a riboflavin kinase homolog in plants
Creators: Francisco J Sandoval - Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164, USA
Sanja Roje
Publication Details: The Journal of biological chemistry, Vol.280(46), pp.38337-38345
Academic Unit: Biological Chemistry, Institute of
Publisher: United States
Identifiers: 99900547473101842
Language: English
Resource Type: Journal article