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Directed immobilization of DNA-binding proteins on a cognate DNA-modified chip surface

Paper ID Volume ID Publish Year Pages File Format Full-Text
24866 43542 2008 6 PDF Available
Title
Directed immobilization of DNA-binding proteins on a cognate DNA-modified chip surface
Abstract

Here we describe a useful method for the site-directed immobilization of proteins with a DNA-binding domain (DNA-BD) on the cognate DNA-coated gold surface for surface plasmon resonance (SPR) imaging analyses. In order to assess the performance of this procedure, we utilized two DNA-BDs, yeast GAL4 DNA-BD, and bacterial LexA DNA-BD. After the immobilization of the cognate double-stranded DNAs (dsDNAs) to a gold chip surface with a monolayer of poly(l-lysine) for sequence-specific DNA–protein interaction, purified recombinant GAL4 DNA-BD:EGFP and LexA DNA-BD:RFP fusion proteins were applied to a dsDNA-spotted gold chip, and were subsequently analyzed using an SPR imaging system. Consequently, the recombinant DNA-binding proteins, GAL4 DNA-BD:EGFP and LexA DNA-BD:RFP, were shown to bind selectively to their cognate DNA sequences on the gold chip. Collectively, our results revealed that sequence-specific dsDNA microarray approach could prove useful in performing the site-directed immobilization of DNA-binding proteins onto a gold thin film in a parallel format, and thereby potentially allowing for the analysis of transcription factor binding profiling as well as for the monitoring of protein–protein interactions between target proteins with DNA-binding domain as a fusion tag and their binding partners.

Keywords
SPR imaging; DNA-binding domain; DNA-modified surface; Site-directed immobilization
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Directed immobilization of DNA-binding proteins on a cognate DNA-modified chip surface
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 135, Issue 1, 20 May 2008, Pages 16–21
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us