Isolation and characterization of two cDNAs encoding 4-coumarate:CoA ligase in Lithospermum cell cultures.
Two near full-length cDNAs (LE4CL-1, LE4CL-2), which encode 4-coumarate:CoA ligase (4CL), were cloned from a library of Lithospermum erythrorhizon cell suspension cultures by the use of heterologous probe of potato 4CL. These cDNAs are 2.1 kb and 2.2 kb in length, respectively. LE4CL-1 encodes 636 amino acids, whose homologies to the 4CL protein sequences known to potato, parsley, pine and rice, were found to be 68%, 66%, 56% and 50% (identities on amino acid level), respectively, whereas those of the predicted translation product of LE4CL-2 (594 amino acids) to the above 4CL proteins were 49 approximately 54%. The similarity of the deduced amino acid sequences between the two 4CLs from Lithospermum cell cultures was 49% in identity. Northern analyses showed that the mRNA levels of both LE4CL-1 and LE4CL-2 were much higher under illumination than in the dark, as reported for the 4CL genes of such plants as parsley. In comparison of mRNA levels of LE4CL-1 and LE4CL-2, the former was demonstrated to be generally higher than the latter by means of an application of RT-PCR. The genomic southern blot experiments suggested that there are probably three copies of LE4CL-1 in the Lithospermum genome DNA, whereas only one copy was detected for LE4CL-2.
Yazaki
K
K
Faculty of Pharmaceutical Sciences, Kyoto University, Japan.
Ogawa
A
A
Tabata
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M
D000595:Amino Acid Sequence; D001483:Base Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003066:Coenzyme A Ligases / Q000145:classification / Q000235:genetics*; D018076:DNA, Complementary; D008969:Molecular Sequence Data; D010802:Phylogeny; D010940:Plant Proteins / Q000235:genetics*; D010944:Plants / Q000201:enzymology / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D018749:RNA, Plant / Q000378:metabolism
D018076:DNA, Complementary; D010940:Plant Proteins; D012333:RNA, Messenger; D018749:RNA, Plant; D003066:Coenzyme A Ligases; C019943:4-coumarate-CoA ligase