M. Frugier et al., A domain in the N-terminal extension of class IIb eukaryotic aminoacyl-tRNA synthetases is important for tRNA binding, EMBO J, 19(10), 2000, pp. 2371-2380
Cytoplasmic aspartyl-tRNA synthetase (AspRS) from Saccharomyces cerevisiae
is a homodimer of 64 kDa subunits, Previous studies have emphasized the hig
h sensitivity of the N-terminal region to proteolytic cleavage, leading to
truncated species that have lost the first 20-70 residues but that retain e
nzymatic activity and dimeric structure, In this work, we demonstrate that
the N-terminal extension in yeast AspRS participates in tRNA binding and we
generalize this finding to eukaryotic class IIb aminoacyl-tRNA synthetases
. By gel retardation studies and footprinting experiments on yeast tRNA(ASp
), We show that the extension, connected to the anticodon-binding module of
the synthetase, contacts tRNA on the minor groove side of its anticodon st
em. Sequence comparison of eukaryotic class IIb synthetases identifies a ly
sine-rich 11 residue sequence ((29)LSKKALKKLQK(39) in yeast AspRS with the
consensus xSKxxLKKxxK in class IIb synthetases) that is important for this
binding. Direct proof of the role of this sequence comes from a mutagenesis
analysis and from binding studies using the isolated peptide.