Monitoring of conformational change in maltose binding protein using split green fluorescent protein = Split GFP assay를 이용한 MBP 구조변화 측정

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Title
Monitoring of conformational change in maltose binding protein using split green fluorescent protein = Split GFP assay를 이용한 MBP 구조변화 측정
Author(s)
Jin Young Jeong; Sang Kyu Kim; Jun Hyoung Ahn; Kyoungsook Park; Eun-Ju Jeong; Moonil Kim; Bong Hyun Chung
Bibliographic Citation
Biochemical and Biophysical Research Communications, vol. 339, no. 2, pp. 647-651
Publication Year
2006
Abstract
In this study, we describe a novel method for the detection of conformational changes in proteins, which is predicated on the reconstitution of split green fluorescent protein (GFP). We employed fluorescence complementation assays for the monitoring of the conformationally altered proteins. In particular, we used maltose binding protein (MBP) as a model protein, as MBP undergoes a characteristic hinge-twist movement upon substrate binding. The common feature of this approach is that GFP, as a reporter protein, splits into two non-fluorescent fragments, which are genetically fused to the N- and C-termini of MBP. Upon binding to maltose, the chromophores move closer together, resulting in the generation of fluorescence. This split GFP method also involves the reconstitution of GFP, which is determined via observations of the degree to which fluorescence intensity is restored. As a result, reconstituted GFP has been observed to generate fluorescence upon maltose binding in vitro, thereby allowing for the direct detection of changes in fluorescence intensity in response to maltose, in a concentration- and time-dependent fashion. Our findings showed that the fluorescence complementation assay can be used to monitor the conformational alterations of a target protein, and this ability may prove useful in a number of scientific and medical applications.
Keyword
Conformational changeGreen fluorescent proteinMaltose binding protein
ISSN
0006-291X
Publisher
Elsevier
DOI
http://dx.doi.org/10.1016/j.bbrc.2005.11.056
Type
Article
Appears in Collections:
Division of Research on National Challenges > Environmental diseases research center > 1. Journal Articles
Critical Diseases Diagnostics Convergence Research Center > 1. Journal Articles
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