Construction of intra-domain chimeras of aminoacyl-tRNA synthetases.
To explore new approaches to enzyme engineering, intra-domain chimeras of two aminoacyl-tRNA synthetases were constructed. Connections were made within the nucleotide folds of these enzymes at sites earlier shown either to be dispensable for activity or able to accommodate oligopeptide insertions. (R.M. Starzyk, T.A. Webster and P. Schimmel, Science 237, 1614 (1987); R.M. Starzyk, J.J. Burbaum and P. Schimmel, Biochemistry, in press). Based on the known structure of one synthetase and structural modeling of the other, the locations of the connection sites allow the possibility of functional "compound" ATP and tRNA binding sites. Of five chimeric genes which were constructed, three direct synthesis of polypeptides that accumulate in vivo. These stable hybrids provide prototypes to which mutagenesis procedures may be applied to produce enzymatically active chimeric synthetases.
Starzyk
R M
RM
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Schimmel
P
P
D000255:Adenosine Triphosphate / Q000378:metabolism; D000604:Amino Acyl-tRNA Synthetases / Q000096:biosynthesis*; D004926:Escherichia coli / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D015202:Protein Engineering; D012355:RNA, Transfer, Ile / Q000378:metabolism; D012358:RNA, Transfer, Met / Q000378:metabolism; D014170:Transformation, Genetic
D012355:RNA, Transfer, Ile; D012358:RNA, Transfer, Met; D000255:Adenosine Triphosphate; D000604:Amino Acyl-tRNA Synthetases