The N-terminal region of DNA polymerase delta catalytic subunit is necessary for holoenzyme function.
Genetic and biochemical studies have shown that DNA polymerase delta (Poldelta) is the major replicative Pol in the eukaryotic cell. Its functional form is the holoenzyme composed of Poldelta, proliferating cell nuclear antigen (PCNA) and replication factor C (RF-C). In this paper, we describe an N-terminal truncated form of DNA polymerase delta (DeltaN Poldelta) from calf thymus. The DeltaN Poldelta was stimulated as the full-length Poldelta by PCNA in a RF-C-independent Poldelta assay. However, when tested for holoenzyme function in a RF-C-dependent Poldelta assay in the presence of RF-C, ATP and replication protein A (RP-A), the DeltaN Poldelta behaved differently. First, the DeltaN Poldelta lacked holoenzyme functions to a great extent. Second, product size analysis and kinetic experiments showed that the holoenzyme containing DeltaN Poldelta was much less efficient and synthesized DNA at a much slower rate than the holoenzyme containing full-length Poldelta. The present study provides the first evidence that the N-terminal part of the large subunit of Poldelta is involved in holo-enzyme function.
Schumacher
S B
SB
Institute of Veterinary Biochemistry, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland.
Stucki
M
M
Hübscher
U
U
D000818:Animals; D020134:Catalytic Domain; D002417:Cattle; D004258:DNA Polymerase III / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D018398:Homeodomain Proteins; D015778:Minor Histocompatibility Antigens; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D051818:Replication Protein C; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins
C096816:BCL2-related protein A1; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; C096063:MATA1 protein, S cerevisiae; D015778:Minor Histocompatibility Antigens; D018809:Proliferating Cell Nuclear Antigen; D019253:Proto-Oncogene Proteins c-bcl-2; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D004258:DNA Polymerase III; D051818:Replication Protein C