title: Zinc-dependent cell growth conferred by mutant tRNA synthetase.
abstract: We present evidence that zinc bound near the C terminus of a long tRNA synthetase polypeptide, and at a location far in the sequence from the catalytic domain, is needed to sustain cell growth and is, therefore, essential for enzyme function. Several class I and class II tRNA synthetases contain bound zinc, including the 939-amino acid class I Escherichia coli isoleucyl-tRNA synthetase, which has two zinc atoms coordinated to cysteine sulfhydryls. The functional significance of these bound zinc atoms has been unclear. Like other class I tRNA synthetases, the isoleucine enzyme has a class-defining conserved N-terminal domain that contains the catalytic site. The C-terminal domain is variable in sequence and structure and not conserved among all of the class I enzymes. Using split proteins, we localized a zinc binding site to the C-terminal end of isoleucyl-tRNA synthetase. Serine substitutions of single cysteines at a thiol-containing putative zinc binding site that is less than 40 amino acids from the C terminus confer a zinc-dependent growth phenotype on cells harboring the mutant enzymes. We propose that zinc bound near the C terminus is part of a structure that interacts directly or indirectly with the active site. A structure at the C terminus that provides a functional link between the conserved N-terminal catalytic and non-conserved C-terminal domain may be common to several class I enzymes.
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
authors.forename: J A
authors.initials: JA
authors.lastname: Landro
authors.forename: P
authors.initials: P
authors.lastname: Schimmel
mesh_terms: D001665:Binding Sites; D002455:Cell Division; D003545:Cysteine / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology; D007533:Isoleucine-tRNA Ligase / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D015032:Zinc / Q000378:metabolism*
chemical_list: D007533:Isoleucine-tRNA Ligase; D015032:Zinc; D003545:Cysteine