Expression, purification and structural analysis of the Pyrococcus abyssi RNA binding protein PAB1135

Here we report the expression, purification and structural analysis of PAB1135. We analyzed the interaction of PAB1135 with RNA and show that it binds efficiently double-stranded RNAs in a non-sequence specific manner. We also performed molecular modeling of the PAB1135 structure using the crystal structure of the protein Af2318 from Archaeoglobus fulgidus (2OGK) as the template.Comparison of this model has lead to the identification of a region in PAB1135 that could be involved in recognizing double-stranded RNA.Despite the recent progress in various genome analysis projects, about a quarter of the archaeal genomes encode functionally uncharacterized proteins, which are almost all only common to other archaeal species [1-4]. Pyrococcus abyssi PAB1135 protein function has not yet been characterized, and it is classified in the family domain of unknown function 54 (DUF54) and in the uncharacterized protein family 0201 (UPF0201). This group's members have been annotated as conserved hypothetical proteins in 46 archaeal species. Some of these proteins were annotated as possible exosome subunits (TK1451 - GI: 57641386, Thermococcus kodakarensis; MK0388 - GI: 20093826, Methanopyrus kandleri; Msp1244 - GI: 84490032, Methanosphaera stadtmanae) [4-6], but analysis of completely sequenced Pyrococcus abyssi genome revealed the presence ofPAB1135 gene in the same operon as Pa1136, the ribonuclease P (RNase P) subunit Rpp30 [7].RNase P is an endoribonuclease responsible for maturation of tRNAs in all domains of life. RNase P is a ribonucleoprotein (RNP) complex, formed by one RNA molecule and a variable number of protein subunits, depending on the organism. Bacterial RNase P contains one protein, whereas the archaeal relative contains at least four proteins, and in humans, it contains at least 10 protein subunits [8,9]. Pyrococcus horikoshii RNase P has been shown to be formed by one catalytic RNA and the proteins Ph1481, Ph1601, Ph1771, and Ph1877, which show homology to the h