Regulation of enzyme activity of alcohol dehydrogenase through its interactions with pyruvate-ferredoxin oxidoreductase in Thermoanaerobacter tengcongensis.
Alcohol dehydrogenases (ADHs) from thermophilic microorganisms are interesting enzymes that have their potential applications in biotechnology and potentially provide insight into the mechanisms of action of thermo-tolerant proteins. The molecular mechanisms of ADHs under thermal stress in vivo have yet to be explored. Herein, we employed a proteomic strategy to survey the possible interactions of secondary-ADH (2-ADH) with other proteins in Thermoanaerobacter tengcongensis (T. tengcongensis) cultured at 75°C and found that 2-ADH, pyruvate-ferredoxin oxidoreductase (PFOR) and several glycolytic enzymes coexisted in a protein complex. Using anion exchange chromatography, the elution profile indicated that the native 2-ADH was present in two forms, PFOR-bound and PFOR-free. Immuno-precipitation and pull down analysis further validated the interactions between 2-ADH and PFOR. The kinetic behaviours of 2-ADH either in the recombinant or native form were evaluated with different substrates. The enzyme activity of 2-ADH was inhibited in a non-competitive mode by PFOR, implying the interaction of 2-ADH and PFOR negatively regulated alcohol formation. In T. tengcongensis, PFOR is an enzyme complex located at the upstream of 2-ADH in the alcohol generation pathway. These findings, therefore, offered a plausible mechanism for how alcohol metabolism is regulated by hetero-interactions between 2-ADH and PFOR, especially in anaerobic thermophiles.
Beijing Institute of Genomics, Chinese Academy of Sciences, China.
Qian
Q
Wang
Quanhui
Q
Wang
Wei
W
Tong
Xue
X
Bai
Zhen
Z
Chen
Jingjing
J
Zhao
Jiyuan
J
Zhang
Siqi
S
Liu
D000426:Alcohol Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D047468:Immunoprecipitation; D007700:Kinetics; D040901:Proteomics; D050743:Pyruvate Synthase / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D046968:Thermoanaerobacter / Q000201:enzymology*
D011994:Recombinant Proteins; D000426:Alcohol Dehydrogenase; D050743:Pyruvate Synthase