Journal
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
Volume 13, Issue 2, Pages 2459-2471Publisher
MDPI
DOI: 10.3390/ijms13022459
Keywords
secondary structure; circular dichroism; membrane protein; organic cosolvent; beta-barrel; FhuA; precipitation; structural integrity
Funding
- Excellence Initiative by the German federal and state governments
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Circular dichroism (CD) and deconvolution were used to study the structural integrity of a plugged and an open FhuA transmembrane channel protein in the presence of varied concentrations of tetrahydrofuran (THF), ethanol (EtOH) and chloroform/methanol (C/M). FhuA is an Escherichia coli outer membrane protein (78.9 kDa) consisting of 22 beta-sheets and an internal globular cork domain which acts as an iron transporter. FhuA and the deletion variant FhuA Delta 1-159 showed comparable and remarkable resistance in the presence of THF (<= 40 vol%) and EtOH (<= 10 vol%). In C/M, significant differences in structural resistance were observed (FhuA stable <= 10 vol%; FhuA Delta 1-159 <= 1 vol%). Deconvolution of CD-spectra for FhuA and FhuA Delta 1-159 yielded beta-sheet contents of 61 % (FhuA) and 58% (FhuA Delta 1-159). Interestingly, FhuA and FhuA Delta 1-159 had comparable beta-sheet contents in the presence and absence of all three organic cosolvents. Additionally, precipitated FhuA and FhuA Delta 1-159 (in 40 vol% C/M or 65 vol% THF) redissolved by supplementing the detergent n-octyl-oligo-oxyethylene (oPOE).
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