혈청 칼슘 농도에 미치는 넙치 유전자 재조합 스타니오칼신의 효과
Effect of Recombinant Olive Flounder Stanniocalcin on Serum Calcium Levels
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Stanniocalcin 1 (STC1) is a glycoprotein hormone that is important in the maintenance of calcium and phosphate homeostasis in both fish and mammals. STC1 and its paralog STC2 are expressed in multiple tissues in fishes, although the physiological roles of piscine STCs are still unclear compared with those of mammals. In this study, we cloned olive flounder STC1 (ofSTC1) and ofSTC2 cDNAs into pET28a vector and used E. coli Rosetta (DE3) as the host strain for protein expression. Expression experiments were carried out using isopropyl-β-D-thiogalactoside (IPTG) and nickel affinity chromatography. We could identify the recombinant proteins as single 29.5 kDa (ofSTC1) and 33.2 kDa (ofSTC2) bands in the insoluble fraction on sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE). These results indicate that ofSTC1 and ofSTC2 were expressed as insoluble proteins in E. coli. Furthermore, the injection of ofSTC1 protein into juvenile tilapia resulted in a decrease of the serum calcium level. These results suggest that the purified fish STC1 and STC2 proteins may be used to elucidate the physiological role of STCs in fishes.
Abstract
Stanniocalcin 1 (STC1) is a glycoprotein hormone that is important in the maintenance of calcium and phosphate homeostasis in both fish and mammals. STC1 and its paralog STC2 are expressed in multiple tissues in fishes, although the physiological roles of piscine STCs are still unclear compared with those of mammals. In this study, we cloned olive flounder STC1 (ofSTC1) and ofSTC2 cDNAs into pET28a vector and used E. coli Rosetta (DE3) as the host strain for protein expression. Expression experiments were carried out using isopropyl-β-D-thiogalactoside (IPTG) and nickel affinity chromatography. We could identify the recombinant proteins as single 29.5 kDa (ofSTC1) and 33.2 kDa (ofSTC2) bands in the insoluble fraction on sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE). These results indicate that ofSTC1 and ofSTC2 were expressed as insoluble proteins in E. coli. Furthermore, the injection of ofSTC1 protein into juvenile tilapia resulted in a decrease of the serum calcium level. These results suggest that the purified fish STC1 and STC2 proteins may be used to elucidate the physiological role of STCs in fishes.
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