Crystallization and preliminary crystallographic studies of putative threonyl-tRNA synthetases fromAeropyrum pernixandSulfolobus tokodaii

Abstract

Threonyl-tRNA synthetase (ThrRS) plays an essential role in protein synthesis by catalyzing the aminoacylation of tRNA(Thr) and editing misacylation. ThrRS generally contains an N-terminal editing domain, a catalytic domain and an anticodon-binding domain. The sequences of the editing domain in ThrRSs from archaea differ from those in bacteria and eukaryotes. Furthermore, several creanarchaea including Aeropyrum pernix K1 and Sulfolobus tokodaii strain 7 contain two genes encoding either the catalytic or the editing domain of ThrRS. To reveal the structural basis for this evolutionary divergence, the two types of ThrRS from the crenarchaea A. pernix and S. tokodaii have been overexpressed in Eschericha coli, purified and crystallized by the hanging-drop vapour-diffusion method. Diffraction data were collected and the structure of a selenomethionine-labelled A. pernix type-1 ThrRS crystal has been solved using the MAD method.

Two Complementary Enzymes for Threonylation of tRNA in Crenarchaeota: Crystal Structure of Aeropyrum pernix Threonyl-tRNA Synthetase Lacking a cis-Editing Domain

“…The sequence of ApThrRS-2 can be divided into three domains: the editing domain, an additional (unknown) domain of 109 residues, and the anticodon-binding domain. 8 The amino acid sequence in the editing domain of ApThrRS-2 is similar to that of PaThrRS with an identity of 37%. It has been reported that the two editing domains of PaThrRS form a dimer.…”

“…Therefore, it is apparent that the editing domains of EcThrRS are completely missing in ApThrRS-1, as predicted from sequence alignments. 8 The N-terminal and the Cterminal domains of ApThrRS-1 could be the catalytic and the anticodon-binding domains, respectively, and such possibilities will be further discussed with the detailed structures.…”

“…6 Its N-terminal domain was identified to have a cisediting function, 7 but it has no sequence homology to those of bacterial ThrRSs. 8 The X-ray structure 7 of the editing domain of PaThrRS is quite different from the corresponding structures of EcThrRS 3 and SaThrRS. 4 There is also a low sequence similarity in the two remaining domains between the archaeal and bacterial ThrRSs.…”

“…This is quite different from the case of the trans-editing ThrRS-ed, which has the ability to recognize anticodons, suggesting a divergence in the evolutionary processes of these two proteins. Our previous study 8 showed that the crenarchaeal Aeropyrum pernix, Sulfolobus tokodaii, Sulfolobus acidocaldarius, and Metallosphaera sedula genomes contain the two different ThrRS genes (ThrRS-1 and ThrRS-2). ApThrRS-1 contains a catalytic domain and an anticodon-binding domain but no editing domain.…”

“…4 There is also a low sequence similarity in the two remaining domains between the archaeal and bacterial ThrRSs. 8 Recently, two different genes have been assigned for ThrRS in the crenarchaeal Sulfolobus solfataricus genome. It has been suggested that one of the encoded enzymes has a catalytic function (ThrRS-cat) and the other has an editing function (ThrRS-ed).…”