Analysis of the structure and function of the S.pombe DNA ligase I protein Cdc17

• Main Author: Martin, Ina Verena
• Published: University of Edinburgh 2002
• Subjects: 572.7
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.657361

DNA ligases join breaks in double-stranded DNA and are therefore crucial enzymes in all aspects of DNA metabolism. Eukaryotic DNA ligase I homologues belong to the family of ATP-dependent ligases and join Okazaki fragments generated on the lagging strand during chromosomal DNA replication. DNA ligase I enzymes consist of C-terminal catalytic domains conserved across all ATP-dependent ligases, a middle conserved domain of unknown function and N-terminal extensions, which share a conserved PCNA binding motif, but otherwise show very limited sequence similarity. In this work structure-function analyses on the DNA ligase I homologue Cdc17 of <i>Schizosaccharomyces pombe </i>were performed. The presence of the middle conserved non-catalytic domain in addition to the catalytic domains was found to be essential for rescue of a <i>cdc17</i> deletion strain. The N-terminal domain targets the enzyme to the nucleus and mitochondria and essential residues were identified which are required for targeting to both cellular compartments. Evidence suggests that mitochondrial Cdc17 function is required for survival of <i>S. pombe</i>. The PCNA binding motif is functional <i>in vivo</i> since its absence reduces cell viability. Despite these identified function overexpression of truncations lacking the N-terminal domain can complement a <i>cdc17</i> deletion strain.