title: An idiosyncratic serine ordering loop in methanogen seryl-tRNA synthetases guides substrates through seryl-tRNASer formation.
abstract: Seryl-tRNA synthetases (SerRS) covalently attach serine to cognate tRNA(Ser). Atypical SerRSs, considerably different from canonical enzymes, have been found in methanogenic archaea. A crystal structure of methanogenic-type SerRS revealed a motif within the active site (serine ordering loop; SOL), which undergoes a notable induced-fit rearrangement during serine binding. The loop rearranges from a disordered conformation in the unliganded enzyme, to an ordered structure comprising an α-helix followed by a loop. We performed kinetic and thermodynamic analyses of SerRS variants to establish the role of the SOL in serylation. Thermodynamic data confirmed a linkage between binding of serine and α-helix formation, previously described by the crystallographic analysis. The ability of the SOL to adopt the observed secondary structure was recognized as essential for serine activation. Mutation of Gln400, which according to the structural data establishes the main connection between the serine and the SOL, produced only modest kinetic effects. Kinetic data offer new insights into the coupling of the conformational change with active site assembly. Productive positioning of the SOL may be driven by the interaction between Trp396 and the serine α-amino group. Rapid kinetics reveals that His250, a non-SOL residue, is essential for transfer of serine to tRNA. Modeling data established that accommodation of the tRNA within the active site may require movement of the SOL. This would enable His250 to assist in productive positioning of the 3'-end of the tRNA for the aminoacyl transfer. Thus, the rearrangements of the SOL conformationally adjust the active site for both reaction steps.
authors.affiliation: Department of Chemistry, Faculty of Science, University of Zagreb, Horvatovac 102a, 10 000 Zagreb, Croatia. mdulic@chem.pmf.hr
authors.forename: Morana
authors.initials: M
authors.lastname: Dulic
authors.forename: Josip
authors.initials: J
authors.lastname: Pozar
authors.forename: Silvija
authors.initials: S
authors.lastname: Bilokapic
authors.forename: Ivana
authors.initials: I
authors.lastname: Weygand-Durasevic
authors.forename: Ita
authors.initials: I
authors.lastname: Gruic-Sovulj
mesh_terms: D001665:Binding Sites; D002151:Calorimetry; D020134:Catalytic Domain; D002852:Chromatography, Ion Exchange; D017433:Protein Structure, Secondary; D012714:Serine-tRNA Ligase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D013816:Thermodynamics
chemical_list: D012714:Serine-tRNA Ligase