Abstract
In Archaea, type IV prepilins and prearchaellins are processed by designated signal peptidase III (SPaseIII) prior to their incorporation into pili and the archaellum, respectively. These peptidases belong to the family of integral membrane aspartic acid proteases that contain two essential aspartate residues of which the second aspartate is located in a conserved GxGD motif. To this group also bacterial type IV prepilin peptidases, Alzheimer disease-related secretases, signal peptide peptidases and signal peptide peptidase-like proteases in humans belong. Here we have performed detailed in vivo analyses to understand the cleavage activity of PibD, SPaseIII from the thermoacidophilic crenarchaeon Sulfolobus acidocaldarius. Using an already established in vivo heterologous system cleavage assay, we could successfully identify the key amino acid residues essential for catalysis of PibD. Furthermore, in trans complementation of a pibD S. acidocaldarius deletion mutant with PibD variants having substituted key amino acids has consolidated our observations of the importance of these residues in catalysis. Based on our data, we propose to re-define class III peptidases/type IV prepilin/prearchaellin peptidases as GxHyD group (rather than GxGD) of proteases [Hy-hydrophobic amino acid].
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Acknowledgments
We are grateful to Dr. Andreas Klingl for assistance in electron microscopy and Prof. Uwe-G. Maier for allocation of the electron microscopic facilities. A.K. was supported by the LOEWE Research Centre for Synthetic Microbiology (Synmikro). A. L. H. was supported by the German Research Council within the framework of the Collaborative Research Center 987 “Microbial Diversity in Environmental Signal Response”. M. v. W. was supported by a grant from the German Research Council AL1206/3-1 and SVA received intramural funds from the Max Planck Society.
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Communicated by M. W. W. Adams.
This article is part of a special issue based on the 10th International Congress on Extremophiles held in Saint Petersburg, Russia, September 7–11, 2014.
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Henche, AL., van Wolferen, M., Ghosh, A. et al. Dissection of key determinants of cleavage activity in signal peptidase III (SPaseIII) PibD. Extremophiles 18, 905–913 (2014). https://doi.org/10.1007/s00792-014-0675-4
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DOI: https://doi.org/10.1007/s00792-014-0675-4