Molecular characterization of a novel protein disulfide isomerase in carrot.
A protein disulfide isomerase (PDI) coding sequence was cloned from a cDNA library derived from carrot (Daucus carota L.) somatic embryos. The cDNA is 2060 bp in length and encodes for a protein of 581 amino acids and molecular weight of 64.4 kDa. Primary structure analysis of the deduced protein revealed two thioredoxin-like active sites and an endoplasmic reticulum-retention signal at its C-terminus, which is also found in PDIs in plants and animals. Although between the carrot protein and other plant PDIs there is only about 30% identity, the active site regions are almost identical. The corresponding mRNA was found in varying amounts, in all tissues investigated. A recombinant protein expressed from the carrot cDNA clone effectively catalyzed both glutathione-insulin transhydrogenation and the oxidative renaturation of denatured RNase A. These results suggest that the protein coded for by the carrot gene is a novel member of the PDI family in plants. We therefore designated this novel carrot gene PDIL1. The protein expressed by the PDIL1 cDNA sequence had a highly acidic stretch at its N-terminal region (no such domain exists in known plant PDIs), and was located far from known plant PDIs on a maximum likelihood tree. The PDIL1 gene, together with closely-related genes identified in Arabidopsis and tomato, was suggested to belong to a novel subfamily of PDIs.
Xu
Zheng-Jun
ZJ
Biotechnology Institute, Akita Prefectural University, Ohgata, Akita 010-0444, Japan.
Ueda
Kenji
K
Masuda
Kiyoshi
K
Ono
Michiyuki
M
Inoue
Masayasu
M
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D018552:Daucus carota / Q000201:enzymology / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D010802:Phylogeny; D019704:Protein Disulfide-Isomerases / Q000235:genetics*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D019704:Protein Disulfide-Isomerases