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Chloroplast Cpn20 forms a tetrameric structure in Arabidopsis thaliana
Author(s) -
Koumoto Yasuko,
Shimada Tomoo,
Kondo Maki,
Takao Toshifumi,
Shimonishi Yasutsugu,
HaraNishimura Ikuko,
Nishimura Mikio
Publication year - 1999
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1046/j.1365-313x.1999.00388.x
Subject(s) - groes , chaperonin , transit peptide , groel , biology , biochemistry , chloroplast , arabidopsis thaliana , homotetramer , complementary dna , escherichia coli , tetramer , arabidopsis , chaperone (clinical) , peptide sequence , microbiology and biotechnology , protein subunit , protein folding , gene , plastid , mutant , medicine , pathology , enzyme
Summary Chloroplast chaperonin 20 (Cpn20) in higher plants is a functional homologue of theEscherichia coliGroES, which is a critical regulator of chaperonin‐mediated protein folding. The cDNA for a Cpn20 homologue ofArabidopsis thalianawas isolated. It was 958 bp long, encoding a protein of 253 amino acids. The protein was composed of an N‐terminal chloroplast transit peptide, and the predicted mature region comprised two distinct GroES domains that showed 42% amino acid identity to each other. The isolated cDNA was constitutively expressed in transgenic tobacco. Immunogold labelling showed that Cpn20 is accumulated in chloroplasts of transgenic tobacco. A Northern blot analysis revealed that mRNA for the chloroplast Cpn20 is abundant in leaves and is increased by heat treatment. To examine the oligomeric structure of Cpn20, a histidine‐tagged construct lacking the transit peptide was expressed inE. coliand purified by affinity chromatography. Gel‐filtration and cross‐linking analyses showed that the expressed products formed a tetramer. The expressed products could substitute for GroES to assist the refolding of citrate synthase under non‐permissive conditions. The analysis on the subunit stoichiometry of the GroEL–Cpn20 complex also revealed that the functional complex is composed of a GroEL tetradecamer and a Cpn20 tetramer.