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Cloning,overexpression and purification of functionally active Saccharomyces cerevisiae Hop1 protein from scherichia coli

Khan, Krishnendu and Madhavan, TP Vipin and Muniyappa, K (2010) Cloning,overexpression and purification of functionally active Saccharomyces cerevisiae Hop1 protein from scherichia coli. In: Protein Expression and Purification, 72 (1). pp. 42-47.

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Official URL: http://dx.doi.org/10.1016/j.pep.2010.03.016

Abstract

One of the major limitations to the application of high-resolution biophysical techniques such as X-crystallography and spectroscopic analyses to structure-function studies of Saccharomyces cerevisiae Hop1 protein has been the non-availability of sufficient quantities of functionally active pure protein. This has, indeed, been the case of many proteins, including yeast synaptonemal complex proteins. In this study, we have performed expression screening in Escherichia coli host strains, capable of high-level expression of soluble S. cerevisiae Hop1 protein. A new protocol has been developed for expression and purification of S. cerevisiae Hop1 protein, based on the presence of hexa-histidine tag and double-stranded DNA-Cellulose chromatography. Recombinant S. cerevisiae Hop1 protein was >98% pure and exhibited DNA-binding activity with high-affinity to the Holliday junction. The availability of the recombinant HOP1 expression vector and active Hop1 protein would facilitate structure-function investigations as well as the generation of appropriate truncated and site-directed mutant proteins, respectively. (C) 2010 Elsevier Inc. All rights reserved.

Item Type: Journal Article
Additional Information: Copyright of this article belongs to Elsevier Science.
Keywords: Meiosis;Synaptonemal complex;Hop1 protein;Holliday junction
Department/Centre: Division of Biological Sciences > Biochemistry
Date Deposited: 02 Jun 2010 05:27
Last Modified: 19 Sep 2010 06:07
URI: http://eprints.iisc.ernet.in/id/eprint/28152

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