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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

Authors 

Jinyoung JeongSang Kyu KimJun Hyoung AhnKyoungsook ParkEun-Ju JeongMoonil KimBong Hyun Chung

Publisher 

Elsevier

Issue Date 

2006

Citation 

Biochemical and Biophysical Research Communications, vol. 339, no. 2, pp. 647-651

Keywords 

conformational changegreen fluorescent proteinmaltose binding proteinmaltosebinding affinityconformational transitionmonitoringcarrier proteinsgreen fluorescent proteinsprotein conformation

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.

ISSN 

0006-291X

Link 

http://dx.doi.org/10.1016/j.bbrc.2005.11.056

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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