In eukaryotes there are three essential DNA polymerases that are involved in the bulk
o f DNA replication: pola, polS and pole. P ola is involved in generating a short
RNA-DNA primer. PolS and 8 are involved in the elongation process o f DNA
replication. It has been suggested that polS is the key enzyme that performs all o f the
processive DNA replication since the catalytic domains o f pole are not essential.In S. pombe polδ is comprised of four subunits: Pol3- the catalytic or A subunit,
Cdcl- the B subunit, Cdc27- the C subunit, and Cdml- the D subunit. Polδ in S.
cerevisiae and mammals have homologues o f these subunits, except for the D
subunit of which there is no homologue in S. cerevisiae.In this thesis polS from S. pombe has been studied in two ways. One approach was to
investigate the protein-protein interactions within polδ, and the other was to
investigate Cdcl, the highly conserved B subunit of unknown function. The proteinprotein interactions were investigated using a combination of two-hybrid assays and
mutational analysis. Cdcl was investigated by performing extensive mutational
analysis using both random and site directed methods.The combination of approaches has demonstrated that the C- terminal ZnF2 region of
both S. pom be and S. cerevisiae A subunit (Pol3) is involved in the direct binding to
the B subunit. The four cysteines present in the zinc finger are involved in
maintaining the structure of both S. pombe and S. cerevisiae ZnF2. Mutational
analysis o f Cdcl (the S. pombe B subunit) has identified a conserved region (DomIII)
that could be involved in the function of Cdcl. Additionally, binding assays with the
Cdcl mutants have suggested a region of Cdcl (from amino acids 293 to 329) as
being involved in the binding to Pol3
