Plant riboflavin biosynthesis. Cloning, chloroplast localization, expression, purification, and partial characterization of spinach lumazine synthase

Lumazine synthase, which catalyzes the penultimate step of riboflavin biosynthesis, has been cloned from three higher plants (spinach, tobacco, and arabidopsis) through functional complementation of an Escherichia coli auxotroph. Whereas the three plant proteins exhibit some structural similarities...

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Bibliographic Details
Published in:The Journal of biological chemistry 1999-07, Vol.274 (31), p.22114-22121
Main Authors: Jordan, D.B, Bacot, K.O, Carlson, T.J, Kessel, M, Viitanen, P.V
Format: Article
Language:English
Subjects:
Amino Acid Sequence
amino acid sequences
Arabidopsis - enzymology
Arabidopsis - genetics
Arabidopsis thaliana
Bacteria - enzymology
Base Sequence
chloroplasts
Chloroplasts - enzymology
Chromatography, Affinity
cloning
Cloning, Molecular
Electrophoresis, Polyacrylamide Gel
enzyme activity
enzymes
Escherichia coli
genbank/af147203
genbank/af148648
genbank/af148649
genes
Genetic Complementation Test
immunocytochemistry
molecular conformation
Molecular Sequence Data
Multienzyme Complexes - chemistry
Multienzyme Complexes - genetics
Multienzyme Complexes - metabolism
Nicotiana - enzymology
Nicotiana - genetics
Nicotiana tabacum
nucleotide sequences
Plants, Toxic
protein transport
Recombinant Proteins - chemistry
Recombinant Proteins - isolation & purification
Recombinant Proteins - metabolism
Riboflavin - biosynthesis
Saccharomyces cerevisiae - enzymology
Sequence Alignment
Sequence Homology, Nucleic Acid
Spinacia oleracea
Spinacia oleracea - enzymology
Spinacia oleracea - genetics
zymogens
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Summary:Lumazine synthase, which catalyzes the penultimate step of riboflavin biosynthesis, has been cloned from three higher plants (spinach, tobacco, and arabidopsis) through functional complementation of an Escherichia coli auxotroph. Whereas the three plant proteins exhibit some structural similarities to known microbial homologs, they uniquely possess N-terminal polypeptide extensions that resemble typical chloroplast transit peptides. In vitro protein import assays with intact chloroplasts and immunolocalization experiments verify that higher plant lumazine synthase is synthesized in the cytosol as a larger molecular weight precursor protein, which is post-translationally imported into chloroplasts where it is proteolytically cleaved to its mature size. The authentic spinach enzyme is estimated to constitute
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.274.31.22114