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The crystal structure of the IleRS and those of the complexes of IleRS with L-isoleucine and L-valine were determined at resolutions of 2.5, 2.8, and 2.8 A, respectively (Table 1). The IleRS structure exhibits the Rossmann-fold domain at the center, ƒÒ-rich intervening domains at the top, and an ƒÑ-rich cylindrical domain at the bottom (Fig. 1).
The Rossmann-fold domain has a central deep catalytic cleft with two characteristic ATP-binding motif. In the L-isoleucineIleRS complex, one isoleucine molecule is bound at the bottom of this catalytic cleft (Fig2, A and B). The hydrophobic side chain of L-isoleucine is recognized by a pocket consisting of Pro46, Trp518, and Trp558 through van der Waals interactions (Fig. 2B). The Asp85 and Gln554 residues form hydrogen bonds with the NH3+ and COO groups, respectively (Fig. 2B). Naturally, larger amino acids are occluded from this pocket. In contrast, in the L-valine¡PIleRS complex structure, an L-valine molecule is actually bound to the same site (Fig. 2C). The hydrophobic contact area of the L-valine side chain with those of Pro46 and Trp558 is slightly smaller than that of the L-isoleucine side chain (Fig. 2, B and C). All of these results agree with the concept of the first, coarse sieve in the double-sieve mechanism of editing.