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ID 39964
file
creator
Nakagawa, Takafumi
Michibata, Hitoshi
subject
Vanadium
Ascidian
Metal-binding Protein
Site-directed mutagenesis
NDC
Biology
abstract
Ascidians are well known to accumulate extremely high levels of vanadium in their blood cells. Several key proteins related to vanadium accumulation and physiological function have been isolated from vanadium-rich ascidians. Of these, vanadium(IV)-binding protein-129 (VBP-129) is a unique protein that has been identified from the blood plasma of an ascidian Ascidia sydneiensis samea, but its metal binding domains are not known. In this study, several deletion and point mutants of VBP-129 were generated, and their metal binding abilities were assessed by immobilized metal ion affinity chromatography (IMAC) and electron spin resonance spectroscopy (ESR). The internal partial protein, VBP-Int41, did not bind to VIV, but the two constructs, VBP-N52 and VBP-Int55, added with additional 11 or 14 neighboring amino acids bound to VIV. Mutations for cysteine-47 and lysine-50 in VBP-Int55 diminished VIV-binding in VBP-Int55, suggesting that these amino acid residues play important roles in binding VIV. ESR titration analysis revealed that VBP-129, VBP-N52 and VBP-Int55 could bind to 6, 3 and 2 VIV ions, respectively. ESR spectrum analysis indicated a N2O2 coordination geometry, which is similar to Vanabins. The cysteines may contribute to the maintenance of the three-dimensional structure that is necessary for binding VIV ions. VBP-129 did not have a VV-reductase activity, as expected from its tissue localization in blood plasma. This study provided the evidences that VBP-129 possesses VIV-binding domains that make a similar coordination to VIV as those by Vanabins but VBP-129 acts solely as VIV-chaperon in blood plasma.
description
This work was supported in part by Grants-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology, Japan (nos. 20570070 and 21570077).
journal title
Journal of Inorganic Biochemistry
volume
Volume 116
start page
70
end page
76
date of issued
2012-11
publisher
Elsevier Inc.
issn
0162-0134
ncid
publisher doi
language
eng
nii type
Journal Article
HU type
Journal Articles
DCMI type
text
format
application/pdf
text version
author
rights
Copyright (c) 2012 Elsevier Inc. All rights reserved.
This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/
relation is version of URL
http://doi.org/10.1016/j.jinorgbio.2012.08.003
department
Graduate School of Science