Molecular cloning and biochemical characterization of a lipoxygenase in almond (Prunus dulcis) seed.
We have characterized an almond (Prunus dulcis) lipoxygenase (LOX) that is expressed early in seed development. The presence of an active lipoxygenase was confirmed by western blot analysis and by measuring the enzymatic activity in microsomal and soluble protein samples purified from almond seeds at this stage of development. The almond lipoxygenase, which had a pH optimum around 6, was identified as a 9-LOX on the basis of the isomers of linoleic acid hydroperoxides produced in the enzymatic reaction. A genomic clone containing a complete lipoxygenase gene was isolated from an almond DNA library. The 6776-bp sequence reported includes an open reading frame of 4667 bp encoding a putative polypeptide of 862 amino acids with a calculated molecular mass of 98.0 kDa and a predicted pI of 5.61. Almond seed lipoxygenase shows 71% identity with an Arabidopsis LOX1 gene and is closely related to tomato fruit and potato tuber lipoxygenases. The sequence of the active site was consistent with the isolated gene encoding a 9-LOX.
Istituto di Ricerca sulle Biotecnologie Agroalimentari, C.N.R, Lecce, Italy. giovanni.mita@irba.le.cnr.it
G
G
Mita
A
A
Gallo
V
V
Greco
C
C
Zasiura
R
R
Casey
G
G
Zacheo
A
A
Santino
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D015342:DNA Probes / Q000235:genetics / Q000378:metabolism; D005091:Exons; D005231:Fatty Acids, Unsaturated / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D006863:Hydrogen-Ion Concentration; D007438:Introns; D008041:Linoleic Acids / Q000378:metabolism; D008054:Lipid Peroxides / Q000378:metabolism; D008084:Lipoxygenase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008861:Microsomes / Q000201:enzymology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D019685:Rosales / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D012639:Seeds / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D012995:Solubility
D015342:DNA Probes; D005231:Fatty Acids, Unsaturated; D008041:Linoleic Acids; D008054:Lipid Peroxides; C008285:linoleic acid hydroperoxide; D008084:Lipoxygenase