期刊
BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES
卷 1818, 期 9, 页码 2202-2208出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbamem.2012.04.015
关键词
AtMRS2; Magnesium transport; Proteoliposome; Arabidopsis thaliana; CorA superfamily
资金
- Ministry of Education, Culture, Sports, Science, and Technology of Japan [22580123]
- Grants-in-Aid for Scientific Research [22580123] Funding Source: KAKEN
Magnesium (Mg2+) plays critical role in many physiological processes. The mechanism of Mg2+ transport has been well documented in bacteria: however, less is known about Mg2+ transporters in eukaryotes. The AtMRS2 family, which consists of 10 Arabidopsis genes, belongs to a eukaryotic subset of the CorA superfamily proteins. Proteins in this superfamily have been identified by a universally conserved GlyMetAsn motif and have been characterized as Mg2+ transporters. Some members of the AtMRS2 family, including AtMRS2-10, may complement bacterial mutants or yeast mutants that lack Mg2+ transport capabilities. Here, we report the purification and functional reconstitution of AtMRS2-10 into liposomes. AtMRS2-10, which contains an N-terminal His-tag, was expressed in Escherichia coli and solubilized with sarcosyl. The purified AtMRS2-10 protein was reconstituted into liposomes. AtMRS2-10 was inserted into liposomes in a unidirectional orientation. Direct measurement of Mg2+ uptake into proteoliposomes revealed that reconstituted AtMRS2-10 transported Mg2+ without any accessory proteins. Mutation in the GMN motif, M400 to I, inactivated Mg2+ uptake. The AtMRS2-10-mediated Mg2+ influx was blocked by Co(III)hexamine, and was independent of the external pH from 5 to 9. The activity of AtMRS2-10 was inhibited by Co2+ and Ni2+; however, it was not inhibited by Ca2+, Fe2+, or Fe3+. While these results indicate that AtMRS2-10 has similar properties to the bacterial CorA proteins, unlike bacterial CorA proteins, AtMRS2-10 was potently inhibited by Al3+. These studies demonstrate the functional capability of the AtMRS2 proteins in proteoliposomes to study' structure-function relationships. (C) 2012 Elsevier B.V. All rights reserved.
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