Isolation and characterization of thioredoxin and NADPH-dependent thioredoxin reductase from tomato (Solanum lycopersicum)

BMB Rep. 2011 Oct;44(10):692-7. doi: 10.5483/BMBRep.2011.44.10.692.

Abstract

To investigate the pathways of oxidoreductases in plants, 2 key components in thioredox systems i.e. thioredoxin h (Trx h) and NADPH-dependent thioredoxin reductase (NTR) genes were first isolated from tomatoes (Solanum lycopersicum). Subsequently, the coding sequences of Trx h and NTR were inserted into pET expression vectors, and overexpressed in Escherichia coli. In the UV-Visible spectra of the purified proteins, tomato Trx h was shown to have a characteristic 'shoulder' at -290 nm, while the NTR protein had the 3 typical peaks unique to flavoenzymes. The activities of both proteins were demonstrated by following insulin reduction, as well as DTNB reduction. Moreover, both NADPH and NADH could serve as substrates in the NTR reduction system, but the catalytic efficiency of NTR with NADPH was 2500-fold higher than with NADH. Additionally, our results reveal that the tomato Trx system might be involved in oxidative stress, but not in cold damage.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Molecular Sequence Data
  • Oxidative Stress
  • Plant Proteins / genetics
  • Plant Proteins / isolation & purification*
  • Plant Proteins / metabolism
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Solanum lycopersicum / chemistry*
  • Solanum lycopersicum / genetics
  • Solanum lycopersicum / metabolism
  • Thioredoxin-Disulfide Reductase / genetics
  • Thioredoxin-Disulfide Reductase / isolation & purification*
  • Thioredoxin-Disulfide Reductase / metabolism
  • Thioredoxins / genetics
  • Thioredoxins / isolation & purification*
  • Thioredoxins / metabolism

Substances

  • Plant Proteins
  • Thioredoxins
  • Thioredoxin-Disulfide Reductase