TY - JOUR
T1 - Cloning, expression, and characterization of cadmium-induced metallothionein-2 from the earthworms Metaphire posthuma and Polypheretima elongata
AU - Liang, Shih Hsiung
AU - Jeng, Yu Ping
AU - Chiu, Yuh Wen
AU - Chen, Jiun Hong
AU - Shieh, Bao Sen
AU - Chen, Chien Yen
AU - Chen, Chien Cheng
N1 - Funding Information:
We want to thank Prof. Michael Yudkin (Oxford University, Oxford, UK) for helpful discussion and Dr. Daniel Chamberlin for critically reading this manuscript. We are grateful to H. J. Chen for technical assistance. This work was supported by a grant from the National Science Council of Taiwan to S.H.L. and C.C.C.
PY - 2009/4
Y1 - 2009/4
N2 - In this study we report the sequences of MT-2 cDNA from two species of Megascoleidae earthworms, Metaphire posthuma and Polypheretima elongata, by mRNA differential display after exposure of the organisms to cadmium. Complementary (c)DNA was verified as the MT-2 gene by the characteristics of its predicted translation product, namely a high cysteine content, conserved CXC motifs, and a molecular weight of around 8 kDa. Amino acid sequence alignment revealed a conserved TKCCG in the cloned MT-2 of both megascolecid earthworms instead of the corresponding conserved TQCCG found in lumbricid earthworms. The cDNAs corresponding to the two megascolecid MT-2 genes were expressed, and the MT-2 proteins were purified for biochemical characterization. The binding of Cu2+ exhibited monophasic kinetics and those of Zn2+ and Cd2+ biphasic kinetics. The proteins bound more tightly to Cd2+ than to Zn2+ and more tightly still to Cu2+. Zn-MT and apo-MT were the most effective at scavenging free radicals, followed by Cd-MT. In conclusion, MT-2s from M. posthuma and P. elongata showed unique sequence features compared to those of lumbricid earthworms. These earthworms could be used to evaluate heavy-metal pollution in soil due to the inducible MT-2 by cadmium exposure.
AB - In this study we report the sequences of MT-2 cDNA from two species of Megascoleidae earthworms, Metaphire posthuma and Polypheretima elongata, by mRNA differential display after exposure of the organisms to cadmium. Complementary (c)DNA was verified as the MT-2 gene by the characteristics of its predicted translation product, namely a high cysteine content, conserved CXC motifs, and a molecular weight of around 8 kDa. Amino acid sequence alignment revealed a conserved TKCCG in the cloned MT-2 of both megascolecid earthworms instead of the corresponding conserved TQCCG found in lumbricid earthworms. The cDNAs corresponding to the two megascolecid MT-2 genes were expressed, and the MT-2 proteins were purified for biochemical characterization. The binding of Cu2+ exhibited monophasic kinetics and those of Zn2+ and Cd2+ biphasic kinetics. The proteins bound more tightly to Cd2+ than to Zn2+ and more tightly still to Cu2+. Zn-MT and apo-MT were the most effective at scavenging free radicals, followed by Cd-MT. In conclusion, MT-2s from M. posthuma and P. elongata showed unique sequence features compared to those of lumbricid earthworms. These earthworms could be used to evaluate heavy-metal pollution in soil due to the inducible MT-2 by cadmium exposure.
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U2 - 10.1016/j.cbpc.2008.09.004
DO - 10.1016/j.cbpc.2008.09.004
M3 - Article
C2 - 18834958
AN - SCOPUS:60949109009
VL - 149
SP - 349
EP - 357
JO - Comparative Biochemistry and Physiology Part - C: Toxicology and Pharmacology
JF - Comparative Biochemistry and Physiology Part - C: Toxicology and Pharmacology
SN - 1532-0456
IS - 3
ER -