Molecular cloning and expression in photosynthetic bacteria of a soybean cDNA coding for phytoene desaturase, an enzyme of the carotenoid biosynthesis pathway.
Carotenoids are orange, yellow, or red photo-protective pigments present in all plastids. The first carotenoid of the pathway is phytoene, a colorless compound that is converted into colored carotenoids through a series of desaturation reactions. Genes coding for carotenoid desaturases have been cloned from microbes but not from plants. We report the cloning of a cDNA for pds1, a soybean (Glycine max) gene that, based on a complementation assay using the photosynthetic bacterium Rhodobacter capsulatus, codes for an enzyme that catalyzes the two desaturation reactions that convert phytoene into zeta-carotene, a yellow carotenoid. The 2281-base-pair cDNA clone analyzed contains an open reading frame with the capacity to code for a 572-residue protein of predicted Mr 63,851. Alignment of the deduced Pds1 peptide sequence with the sequences of fungal and bacterial carotenoid desaturases revealed conservation of several amino acid residues, including a dinucleotide-binding motif that could mediate binding to FAD. The Pds1 protein is synthesized in vitro as a precursor that, upon import into isolated chloroplasts, is processed to a smaller mature form. Hybridization of the pds1 cDNA to genomic blots indicated that this gene is a member of a low-copy-number gene family. One of these loci was genetically mapped using restriction fragment length polymorphisms between Glycine max and Glycine soja. We conclude that pds1 is a nuclear gene encoding a phytoene desaturase enzyme that, as its microbial counterparts, contains sequence motifs characteristic of flavoproteins.
Bartley
G E
GE
Du Pont Central Research and Development, Wilmington, DE 19880-0402.
Viitanen
P V
PV
Pecker
I
I
Chamovitz
D
D
Hirschberg
J
J
Scolnik
P A
PA
D000595:Amino Acid Sequence; D001483:Base Sequence; D002338:Carotenoids / Q000096:biosynthesis*; D002736:Chloroplasts / Q000201:enzymology; D003001:Cloning, Molecular / Q000379:methods; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D010088:Oxidoreductases / Q000235:genetics*; D015183:Restriction Mapping; D016354:Rhodobacter capsulatus / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D013025:Glycine max / Q000201:enzymology / Q000235:genetics*
D015345:Oligonucleotide Probes; D002338:Carotenoids; D004247:DNA; D010088:Oxidoreductases; C022398:phytoene dehydrogenase