AminoacyltRNA Synthetases

Despite the universality of the main features of the three-dimensional structure of tRNAs, aminoacyl-tRNA synthetases (ARSases) show marked differences depending on their amino acid specificity. As a rule, ARSases are relatively large proteins with a molecular mass around 100,000 daltons, although both smaller (about 50 kDa for bacterial CysRS and GluRS) and larger (above 200 kDa for GlyRS, AlaRS and PheRS) enzymes also occur. One third of ARSases are monomeric, half of ARSases are homodimers (a2 type), and the three large ARSases mentioned above are tetramers of a4 or a2ß2 type (see Table 3.1). The molecular masses of subunits of dimeric and tetrameric enzymes range from 35,000 to 90,000 daltons, often being about 50,000 daltons. In the cases of large monomeric enzymes, such as bacterial and fungal

ValRS, LeuRS and IleRS with a molecular mass of 100,000 to 120,000 daltons, it appears that their single polypeptide chain consists of two homologous regions forming two similar domains, each with a molecular mass of about 50,000 to 60,000 daltons. At the same time, ArgRS, CysRS, GluRS and GlnRS of bacteria consist of a single polypeptide chain (also a1 type) with a molecular mass of 50,000 to 60,000 daltons, and do not appear to be subdivided into two homologous regions.

From the analysis of subunit and domain structure of ARSases it is tempting to suggest a generalized pattern of their principal organization. Indeed, most synthetases have a molecular mass around 100,000 daltons and consist either of two subunits or two similar halves (superdomains). Therefore, the principal building unit, i.e. subunit or superdomain, has a molecular mass ranging mainly between 40,000 and 60,000 daltons, and many of the aminoacyl-tRNA synthetases would be considered as dimers or pseudo-dimers of the building unit, i.e. (40,000 - 60,000)2 . The synthetases with greater molecular masses, around 200,000 daltons, may be "duplicated" enzymes of this type. In reality, however, the nonrepeating unit in some cases can be markedly larger; bacterial AlaRS, for example, consists of four identical subunits, each with a molecular mass of about 100,000 daltons, and shows no evidence of any repeats in its amino acid sequence. On the other hand, such relatively small, one-subunit enzymes as CysRS and ArgRS (molecular mass 52,000 and 64,500 daltons, respectively) do not display two homologous superdomains in their structure.

In any case, according to functional tests the molecules of some (but not all) ARSases possess two sets of substrate-binding sites; in other words, they are dimers in the functional sense as well. The active sites, however, are not independent and can markedly affect each other in the dimeric or two-superdomain enzymes, thus displaying a certain cooperativity (see below).

Despite the apparent structural diversity of ARSases, they have been found to possess structural motifs that provide the basis for revealing homology between some of them and for unification of homologous species into classes (Eriani et al., 1990). There are two main distinct classes of ARSases, each including 10 enzymes (Table 3.1). Class I consists of Arg-, Cys-, Gln-, Glu-, Ile-, Leu-, Met-, Trp-, Tyr-, and Val-RSases. They are predominantly monomers, with the exception of Trp- and Tyr-RSases which are homodimers. The monomeric globule is subdivided into different domains. The N-terminal region of the molecule is responsible for the binding of all three substrates, namely ATP, amino acid and the acceptor stem of tRNA, and for the catalysis of the reactions between them. This region is characterized by the

Table 3.1. Classification of E. coli aminoacyl-tRNA synthetases.
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