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Monitoring the activity of single translocons
Taufik I.b, Kedrov A., Exterkate M., Driessen A.J.M.
a Department of Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Netherlands
b School of Life Sciences and Technology, Institut Teknologi Bandung, Indonesia
[vc_row][vc_column][vc_row_inner][vc_column_inner][vc_separator css=”.vc_custom_1624529070653{padding-top: 30px !important;padding-bottom: 30px !important;}”][/vc_column_inner][/vc_row_inner][vc_row_inner layout=”boxed”][vc_column_inner width=”3/4″ css=”.vc_custom_1624695412187{border-right-width: 1px !important;border-right-color: #dddddd !important;border-right-style: solid !important;border-radius: 1px !important;}”][vc_empty_space][megatron_heading title=”Abstract” size=”size-sm” text_align=”text-left”][vc_column_text]Recent studies introduced a novel view that the SecYEG translocon functions as a monomer and interacts with the dimeric SecA ATPase, which fuels the preprotein translocation reaction. Here, we used nanodisc-reconstituted SecYEG to characterize the functional properties of single copies of the translocon. Using a method based on intermolecular Förster resonance energy transfer, we show for the first time that isolated nanodisc-reconstituted SecYEG monomers support preprotein translocation. When several copies of SecYEG were co-reconstituted within a nanodisc, no change in translocation kinetics was observed, suggesting that SecYEG oligomers do not facilitate enhanced translocation. In contrast, nanodisc-reconstituted monomers of the PrlA4 variant of SecYEG showed increased translocation rates. Experiments based on intramolecular Förster resonance energy transfer within the nanodisc-isolated monomeric SecYEG demonstrated a nucleotide-dependent opening of the channel upon interaction with SecA. In conclusion, the nanodisc-reconstituted SecYEG monomers are functional for preprotein translocation and provide a new prospect for single-molecule analysis of dynamic aspects of protein translocation. © 2013. Published by Elsevier Ltd. All rights reserved.[/vc_column_text][vc_empty_space][vc_separator css=”.vc_custom_1624528584150{padding-top: 25px !important;padding-bottom: 25px !important;}”][vc_empty_space][megatron_heading title=”Author keywords” size=”size-sm” text_align=”text-left”][vc_column_text][/vc_column_text][vc_empty_space][vc_separator css=”.vc_custom_1624528584150{padding-top: 25px !important;padding-bottom: 25px !important;}”][vc_empty_space][megatron_heading title=”Indexed keywords” size=”size-sm” text_align=”text-left”][vc_column_text]membrane proteins,protein dynamics,protein oligomerization,protein transport,secretion[/vc_column_text][vc_empty_space][vc_separator css=”.vc_custom_1624528584150{padding-top: 25px !important;padding-bottom: 25px !important;}”][vc_empty_space][megatron_heading title=”Funding details” size=”size-sm” text_align=”text-left”][vc_column_text]We would like to thank Prof. Stephan Sligar for a gift of MSP expression plasmid, Natalya Dudkina for negative-stain electron microscopy experiments, and Daniel N. Wilson for critical reading of the manuscript. This work was financially supported by The Netherlands Foundation for Scientific Research, Chemical Sciences and Earth and Life Sciences . I.T. is a recipient of a scholarship from the Ministry of National Education of the Republic of Indonesia.[/vc_column_text][vc_empty_space][vc_separator css=”.vc_custom_1624528584150{padding-top: 25px !important;padding-bottom: 25px !important;}”][vc_empty_space][megatron_heading title=”DOI” size=”size-sm” text_align=”text-left”][vc_column_text]https://doi.org/10.1016/j.jmb.2013.08.012[/vc_column_text][/vc_column_inner][vc_column_inner width=”1/4″][vc_column_text]Widget Plumx[/vc_column_text][/vc_column_inner][/vc_row_inner][/vc_column][/vc_row][vc_row][vc_column][vc_separator css=”.vc_custom_1624528584150{padding-top: 25px !important;padding-bottom: 25px !important;}”][/vc_column][/vc_row]