Conserved protein TTHA1554 from Thermus thermophilus HB8 binds to glutamine synthetase and cystathionine beta-lyase.
TTHA1554 was found as a hypothetical protein composed of 95 amino acids in the genome of the extremely thermophilic bacterium, Thermus thermophilus HB8. Proteins homologous to TTHA1554 are conserved in several bacteria and archaea, although their functions are unknown. To investigate the function of TTHA1554, we identified interacting proteins by using a pull-down assay and mass spectrometry. TTHA1329, which is glutamine synthetase, and TTHA1620, a putative aminotransferase, were identified as TTHA1554 binding proteins. The interactions with TTHA1329 and TTHA1620 were validated using in vitro pull-down assays and surface plasmon resonance biosensor assays with recombinant proteins. Since sequence homology analyses suggested that TTHA1620 was a pyridoxal 5'-phosphate-dependent enzyme, such as an aminotransferase, a cystathionine beta-lyase or a cystalysin, putative substrates were investigated. When cystathionine, cystine and S-methylcysteine were used as substrates, pyruvate was produced by TTHA1620. The data revealed that TTHA1620 has cystathionine beta-lyase enzymatic activity. When TTHA1554 was added to the reaction mixtures, the glutamine synthetase and cystathionine beta-lyase enzymatic activities both increased by approximately two-fold. These results indicated that TTHA1554 is a novel protein (we named it GCBP: glutamine synthetase and cystathionine beta-lyase binding protein) that binds to glutamine synthetase and cystathionine beta-lyase.
Arai
Ryoichi
R
Protein Research Group, RIKEN Genomic Sciences Center, 1-7-22, Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan.
Nishimoto
Madoka
M
Toyama
Mitsutoshi
M
Terada
Takaho
T
Kuramitsu
Seiki
S
Shirouzu
Mikako
M
Yokoyama
Shigeyuki
S
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D017124:Conserved Sequence; D005974:Glutamate-Ammonia Ligase / Q000378:metabolism*; D008190:Lyases / Q000378:metabolism*; D013058:Mass Spectrometry / Q000379:methods; D025941:Protein Interaction Mapping / Q000379:methods; D016963:Thermus thermophilus / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D008190:Lyases; C019965:cystathionine beta-lyase; D005974:Glutamate-Ammonia Ligase