A Method for Isolation of DNA-Binding Proteins Based on Solubility of DNA-Protein Complexes

Hua      Yang; Huang      Li; Li-qun      Rao; Gui-you      Long; Guo-ping      Peng; Lu      Jin

Abstract

The study of DNA-binding proteins is crucial in understanding gene regulatory networks. We developed a new method for the enrichment of DNA-binding proteins based on the variability of DNA-protein complexes’ solubility in different ionic strength solutions. 0.14M sodium chloride was determined as the most efficient extraction concentration to precipitate DNA-binding proteins. SDS-PAGE analysis revealed that some high-abundance proteins were removed effectively and at the same time DNA-binding proteins were isolated in this simple process. Twenty kinds of proteins were identified in the acquired sample by 1-D gel-LC-MS/MS. Furthermore, computerized analysis of MS data showed that quite a number of unmatched peptides have the classic structure of leucine zipper or zinc finger, which were symbolic elements of transcription factors. These results suggested that this new method can acquire DNA-binding proteins effectively and allow improvement in the isolation of high-quality DNA-binding proteins.

Keywords: DNA-binding protein, DNA-protein Complexes, isolation, solubility, gene regulatory networks, SDS-PAGE, zinc finger, transcription factors