Outside the unusual cell wall of the hyperthermophilic archaeon Aeropyrum pernix K1

In contrast to the extensively studied eukaryal and bacterial protein secretion systems, comparatively less is known about how and which proteins cross the archaeal cell membrane. In order to identify secreted proteins of the hyperthermophilic archaeon Aeropyrum pernix K1 we used a proteomic approach to analyze the extracellular and cell surface protein fractions. The experimentally obtained data comprising 107 proteins were compared to the in silico predicted secretome. Due to the lack of signal peptide and cellular localization prediction tools specific for archaeal species, programs trained on eukaryotic and/or gram-positive and gram-negative bacterial signal peptide data sets were used. PSortB gram-negative and gram-positive analysis predicted 21 (1.2% of ORF) and 24 (1.4% of ORF) secreted proteins, respectively from the entire A. pernix K1 proteome, twelve of which were experimentally identified in this work. Six additional proteins were predicted to follow non classical secretion mechanisms using SecP algorithms. According to at least one of the two PSortB predictions 48 proteins identified in the two fractions possess an unknown localization site. In addition, more than half of the proteins do not contain signal peptides recognized by current prediction programs. This suggests that known mechanisms only partly describe archaeal protein secretion. The most striking characteristic of the secretome was the high number of transport-related proteins identified from the ATP-binding cassette (ABC), tripartite ATP-independent periplasmic (TRAP), ATPase, Small Conductance Mechanosensitive Ion Channel (MscS) and Dicarboxylate Amino acid:Cation Symporter (DAACS) transporter families. In particular, identification of 21 solute-binding receptors of the ABC superfamily out of the 24 predicted in silico, confirms that ABC-mediated transport represents the most frequent strategy adopted by A. pernix for solute translocation across the cell membrane.