Thermodynamic properties distinguish human mitochondrial aspartyl-tRNA synthetase from bacterial homolog with same 3D architecture

Abstract

In the mammalian mitochondrial translation apparatus, the proteins and their partner RNAs are coded by two genomes. The proteins are nuclear-encoded and resemble their homologs, whereas the RNAs coming from the rapidly evolving mitochondrial genome have lost critical structural information. This raises the question of molecular adaptation of these proteins to their peculiar partner RNAs. The crystal structure of the homodimeric bacterial-type human mitochondrial aspartyl-tRNA synthetase (DRS) confirmed a 3D architecture close to that of Escherichia coli DRS. However, the mitochondrial enzyme distinguishes by an enlarged catalytic groove, a more electropositive surface potential and an alternate interaction network at the subunits interface. It also presented a thermal stability reduced by as much as 12°C. Isothermal titration calorimetry analyses revealed that the affinity of the mitochondrial enzyme for cognate and non-cognate tRNAs is one order of magnitude higher, but with different enthalpy and entropy contributions. They further indicated that both enzymes bind an adenylate analog by a cooperative allosteric mechanism with different thermodynamic contributions. The larger flexibility of the mitochondrial synthetase with respect to the bacterial enzyme, in combination with a preserved architecture, may represent an evolutionary process, allowing nuclear-encoded proteins to cooperate with degenerated organelle RNAs.

Translation in Mammalian Mitochondria: Order and Disorder Linked to tRNAs and Aminoacyl-tRNA Synthetases

“…3.2 with three of the four crystal structures that have been determined so far for mt-aaRSs for exclusive mitochondrial location (additional crystallographic structures are available for human GlyRS (Cader et al 2007;Xie et al 2006) and for LysRS (Guo et al 2008), aaRSs of dual cytosolic and mitochondrial location). Bovine mt-SerRS, human mt-TyrRS, and human mt-AspRS show an overall architecture close to that of their prokaryotic homologs (Bonnefond et al 2007;Chimnaronk et al 2005;Neuenfeldt et al 2013). Mt-PheRS is again the exception.…”

“…For instance, mt-PheRS undergoes a large movement of its anticodonbinding domain upon tRNA binding, switching from a closed to an open conformation (Klipcan et al 2012). The higher thermal sensitivity of human mt-AspRS as compared E. coli AspRS, its more open catalytic groove in the absence of tRNA and the amplitude of thermodynamic terms associated with tRNA binding are also in favor of a more dynamic structure (Neuenfeldt et al 2013). Altogether, Fig.…”

“…The redesigned protein was highly soluble, monodisperse and functionally active in tRNA aminoacylation. It yielded crystals that were suitable for structure determination Neuenfeldt et al 2013). These results suggest that additional criteria should be taken into account for the prediction of the correct MTS cleavage sites and that the definition of the precise N-terminus of mature mt-aaRS should be determined experimentally.…”

“…Positively charged patches at the surface of mt-SerRS were redistributed to bind tRNAs lacking an extended variable region, the hallmark and major identity element of prokaryotic, eukaryotic, and archaeal tRNAs Ser (Chimnaronk et al 2005). The three other mt-aaRSs display a more electropositive tRNA-binding interface, which may favor interactions with the sugar-phosphate backbone of the substrate to compensate for a reduction of specific contacts with identity elements (Neuenfeldt et al 2013). The ability of mt-TyrRS, mt-PheRS, and mt-AspRS to aminoacylate heterologuous tRNAs (Bonnefond et al 2005a;Klipcan et al 2012;Neuenfeldt et al 2013) indicates a much higher substrate tolerance, which may be linked to an increased structural plasticity.…”

“…The three other mt-aaRSs display a more electropositive tRNA-binding interface, which may favor interactions with the sugar-phosphate backbone of the substrate to compensate for a reduction of specific contacts with identity elements (Neuenfeldt et al 2013). The ability of mt-TyrRS, mt-PheRS, and mt-AspRS to aminoacylate heterologuous tRNAs (Bonnefond et al 2005a;Klipcan et al 2012;Neuenfeldt et al 2013) indicates a much higher substrate tolerance, which may be linked to an increased structural plasticity. For instance, mt-PheRS undergoes a large movement of its anticodonbinding domain upon tRNA binding, switching from a closed to an open conformation (Klipcan et al 2012).…”