Molecular characterization of low-temperature-inducible NTR-C in Chlorella vulgaris.
We isolated a cDNA corresponding to a chloroplast NADPH-dependent thioredoxin reductase gene (NTR-C), in Chlorella that is low-temperature-inducible. The obtained cDNA was 1,838 bp in length and coded for 529 amino acids. The deduced amino acid sequence showed higher homology to those of Arabidopsis and rice NTR-C, containing a thioredoxin (Trx) and a thioredoxin reductase (TR), than those of NTR-A (mitochondrial) and NTR-B (cytosolic) from various organisms, which contain only a TR domain and differ in subcellular localization. The results of enzyme assays of partially-purified mature NTR-C protein (mNTR-C), expressed in Escherichia coli with a pET-29b(+) expression vector, provided evidence that the gene included both regions. Northern blot analysis showed a remarkable increase in transcripts under low temperature, while the protein level did not significantly change when examined by using Western blotting with anti-mNTR-C antibodies. The TR activity dependent on NADPH was not enhanced by low temperature despite the substantial increase in transcripts. Based on the results of measurement of peroxiredoxin (Prx) activity and Western blotting using both an extract of Chlorella and purified mNTR-C, the Chlorella was suggested to possess a Prx that interacts with NTR-C.
Machida
Takeshi
T
Department of Bioscience and Biotechnology, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, Fukuoka 812-8581, Japan. be204093machida@yahoo.co.jp
Kato
Eri
E
Ishibashi
Akiko
A
Ohashi
Naoto
N
Honjoh
Ken-Ichi
K
Miyamoto
Takahisa
T
D020418:Algal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D048408:Chlorella vulgaris / Q000201:enzymology*; D054464:Peroxiredoxins / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D013880:Thioredoxin-Disulfide Reductase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D020418:Algal Proteins; D012333:RNA, Messenger; D054464:Peroxiredoxins; D013880:Thioredoxin-Disulfide Reductase