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projects: chromatin enzymes
projects: chromaint factors

projects: DNA ladders
projects: polycistronic expression
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gallery: amino acids
gallery: secondary structure
gallery: selected proteins
gallery: protein/DNA complexes

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polycistronic expression vector   coexpression of VCB complex

A major challenge facing structural biologists interested in multicomponent protein complexes is how to isolate milligram quantities of functional protein complexes. In vitro reconstitution of individually expressed and purified protein components is a common, but time-consuming and inefficient process. We are studying in vivo reconstitution by polycistronic expression in E. coli as an alternative to in vitro reconstitution.

We have developed a modular polycistronic expression system for producing protein complexes in E. coli. The first generation pST39 polycistronic expression system enables the coexpression of up to 4 protein subunits from a single plasmid. We have used this polycistronic expression system to produce milligrams of functionally active yeast and human protein complexes containing 2, 3 and 4 subunits, with individual subunit molecular weights of 10-80 kDa.

The second generation pST44 polycistronic expression system improves the modularity of the system and includes a library of affinity tags for efficient purification of the recombinant complexes.

The pST39 and pST44 polycistronic expression system plasmids are available from Addgene:
https://www.addgene.org/Song_Tan/ .

You can download the plasmids maps as a pdf document (650 KB) and the plasmid sequences as a txt file (246 KB).

Present studies are directed at increasing the number of coexpressed subunits and increasing the flexibility of the system.

Reference:

Selleck, W. and S. Tan (2008) Recombinant protein complex expression in E. coli, Curr Protocols Protein Sci, Chapter 5, unit 5.21. (abstract)

Barrios, A., Selleck, W., Hnatkovich. B., Kramer, R., Sermwittayawong, D. and S. Tan (2007) Expression and purification of recombinant yeast Ada2/Ada3/Gcn5 and Piccolo NuA4 histone acetyltransferase complexes, Methods, 41:271-277. (abstract)

Tan, S., Kern, R.C. and Selleck, W. (2005) The pST44 polycistronic expression system for producing protein complexes in Escherichia coli, Prot. Expr. Purif., 40:385-395. (abstract)

Lichty, J.L., Malecki, J.L., Agnew, H.D., Michelson-Horowitz, D.J. and Tan, S. (2005) Comparison of Affinity Tags for Protein Purification, Prot. Expr. Purif., 41:98-105. (abstract)

Tan, S. (2001)A Modular Polycistronic Expression System for Overexpressing Protein Complexes in Escherichia Coli, Prot. Expr. Purif., 21: 224-234. (abstract)


video introduction | projects: | chromatin enzymes | chromatin factors | DNA ladders | polycistronic expression | picture & movie gallery: | amino acids | secondary structure | selected proteins | protein/DNA complexes |
background reading | reprints | positions |
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This page is maintained by Song Tan: sxt30@psu.edu (814-865-3355)
Department of Biochemistry & Molecular Biology, 108 Althouse Lab, University Park, PA 16802

This page was last updated on October 1, 2018.

Penn State BMB Eberly College of Science