Sulfide:quinone oxidoreductase from echiuran worm Urechis unicinctus.
Sulfide is a natural, widely distributed, poisonous substance, and sulfide:quinone oxidoreductase (SQR) has been identified to be responsible for the initial oxidation of sulfide in mitochondria. In this study, full-length SQR cDNA was cloned from the echiuran worm Urechis unicinctus, a benthic organism living in marine sediments. The protein consisted of 451 amino acids with a theoretical pI of 8.98 and molecular weight of 50.5 kDa. Subsequently, the SQR mRNA expression in different tissues was assessed by real-time reverse transcription and polymerase chain reaction and showed that the highest expression was in midgut, followed by anal sacs and coelomic fluid cells, and then body wall and hindgut. Furthermore, activated SQR was obtained by dilution refolding of recombinant SQR expression in E. coli, and the refolded product showed optimal activity at 37 °C and pH 8.5 and K (m) for ubiquinone and sulfide at 15.6 µM and 40.3 µM, respectively. EDTA and GSH had an activating effect on refolded SQR, while Zn(2+) caused decreased activity. Western blot showed that SQR in vivo was located in mitochondria and was ∼ 10 kDa heavier than the recombinant protein. In addition, SQR, detected by immunohistochemistry, was mainly located in the epithelium of all tissues examined. Ultrastructural observations of these tissues' epithelium by transmission electron microscopy provided indirect cytological evidence for its mitochondrial location. Interesting aspects of the U. unicinctus SQR amino acid sequence, its catalytic mechanism, and the different roles of these tissues in sulfide metabolic adaptation are also discussed.
Ma
Yu-Bin
YB
Key Laboratory of Marine Genetics and Breeding, Ministry of Education, Ocean University of China, Qingdao, 266003, China.
Zhang
Zhi-Feng
ZF
Shao
Ming-Yu
MY
Kang
Kyoung-Ho
KH
Tan
Zhi
Z
Li
Jin-Long
JL
D000595:Amino Acid Sequence; D000818:Animals; D000845:Annelida / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D008969:Molecular Sequence Data; D011808:Quinone Reductases / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism
D012333:RNA, Messenger; D011808:Quinone Reductases; C073412:sulfide quinone reductase