title: Analysis of Arabidopsis thioredoxin-h isotypes identifies discrete domains that confer specific structural and functional properties.
abstract: Multiple isoforms of Arabidopsis thaliana h-type thioredoxins (AtTrx-hs) have distinct structural and functional specificities. AtTrx-h3 acts as both a disulfide reductase and as a molecular chaperone. We prepared five representative AtTrx-hs and compared their protein structures and disulfide reductase and molecular chaperone activities. AtTrx-h2 with an N-terminal extension exhibited distinct functional properties with respect to other AtTrx-hs. AtTrx-h2 formed low-molecular-mass structures and exhibited only disulfide reductase activity, whereas the other AtTrx-h isoforms formed high-molecular-mass complexes and displayed both disulfide reductase and molecular chaperone activities. The domains that determine the unique structural and functional properties of each AtTrx-hs protein were determined by constructing a domain-swap between the N- and C-terminal regions of AtTrx-h2 and AtTrx-h3 (designated AtTrx-h-2N3C and AtTrx-h-3N2C respectively), an N-terminal deletion mutant of AtTrx-h2 [AtTrx-h2-N(∆19)] and site-directed mutagenesis of AtTrx-h3. AtTrx-h2-N(∆19) and AtTrx-h-3N2C exhibited similar properties to those of AtTrx-h2, but AtTrx-h-2N3C behaved more like AtTrx-h3, suggesting that the structural and functional specificities of AtTrx-hs are determined by their C-terminal regions. Hydrophobicity profiling and molecular modelling revealed that Ala100 and Ala106 in AtTrx-h3 play critical roles in its structural and functional regulation. When these two residues in AtTrx-h3 were replaced with lysine, AtTrx-h3 functioned like AtTrx-h2. The chaperone function of AtTrx-hs conferred enhanced heat-shock-resistance on a thermosensitive trx1/2-null yeast mutant.
authors.affiliation: *Division of Applied Life Science, PMBBRC, Gyeongsang National University, Jinju 660-701, Republic of Korea.
authors.forename: Young Jun
authors.initials: YJ
authors.lastname: Jung
authors.forename: Yong Hun
authors.initials: YH
authors.lastname: Chi
authors.forename: Ho Byoung
authors.initials: HB
authors.lastname: Chae
authors.forename: Mi Rim
authors.initials: MR
authors.lastname: Shin
authors.forename: Eun Seon
authors.initials: ES
authors.lastname: Lee
authors.forename: Joon-Yung
authors.initials: JY
authors.lastname: Cha
authors.forename: Seol Ki
authors.initials: SK
authors.lastname: Paeng
authors.forename: Yuno
authors.initials: Y
authors.lastname: Lee
authors.forename: Jin Ho
authors.initials: JH
authors.lastname: Park
authors.forename: Woe Yeon
authors.initials: WY
authors.lastname: Kim
authors.forename: Chang Ho
authors.initials: CH
authors.lastname: Kang
authors.forename: Kyun Oh
authors.initials: KO
authors.lastname: Lee
authors.forename: Keun Woo
authors.initials: KW
authors.lastname: Lee
authors.forename: Dae-Jin
authors.initials: DJ
authors.lastname: Yun
authors.forename: Sang Yeol
authors.initials: SY
authors.lastname: Lee
mesh_terms: D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018869:Heat-Shock Response; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics; D009154:Mutation; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry / Q000235:genetics; D055503:Protein Multimerization; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D013696:Temperature; D054480:Thioredoxin h / Q000737:chemistry* / Q000235:genetics
chemical_list: D029681:Arabidopsis Proteins; D018832:Molecular Chaperones; D011994:Recombinant Proteins; D054480:Thioredoxin h; D009247:NADH, NADPH Oxidoreductases