Functional expression and characterization of a bacterial light-harvesting membrane protein in Escherichia coli and cell-free synthesis systems

Bioscience, Biotechnology, and Biochemistry
Yuichiro ShimadaTsunenori Nozawa

Abstract

Heterologous expression of a bacterial light-harvesting (LH) integral membrane protein was attempted using Escherichia coli cells and cell-free synthesis systems prepared from E. coli extracts. The alpha-apoprotein of LH1 complex from purple photosynthetic bacterium Rhodospirillum rubrum was overexpressed as a recombinant protein with a histidine (His6) tag added to the carboxyl terminus. Both of the expression systems produced alpha-apoprotein in a fully functional form as can judged by its ability to form a structural subunit with native beta-apoprotein and the pigment molecule bacteriochlorophyll a. The expression product in E. coli appears to be located in the inner cell membrane and can be almost completely extracted by 0.5% (w/v) Triton X-100. Circular dichroism measurement indicated that the expressed alpha-apoproteins from both systems had alpha-helical contents essentially identical with that of the native one. About two thirds of the alpha-apoprotein expressed in E. coli was found to have the amino terminal methionine residue modified by a formyl group. About one third of the alpha-apoprotein expressed in the cell-free system was found to be oxidized at the side chain of the amino terminal methionine residue. Function...Continue Reading

References

Dec 13, 2003·Science·Aleksander W RoszakRichard J Cogdell

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Citations

Aug 16, 2011·Photosynthesis Research·Anjali Pandit, Huub J M de Groot
Jul 27, 2010·New Biotechnology·Sundaresan RajeshMichael Overduin
Oct 28, 2008·Trends in Biotechnology·Ethan T Johnson, Claudia Schmidt-Dannert
Sep 4, 2008·Proteomics·Daniel SchwarzFrank Bernhard
Jun 7, 2006·Photochemistry and Photobiology·Shiho OnoderaZheng-Yu Wang
Nov 2, 2005·Analytical Biochemistry·Hiroaki SuzukiZheng-Yu Wang
Jul 11, 2006·Magnetic Resonance in Chemistry : MRC·Alexander KoglinVolker Dötsch
Jun 12, 2009·Applied Microbiology and Biotechnology·Jiazhang LianZhinan Xu

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