Send to

Choose Destination
Biochemistry. 2005 Feb 22;44(7):2305-18.

Interaction between the T4 helicase-loading protein (gp59) and the DNA polymerase (gp43): a locking mechanism to delay replication during replisome assembly.

Author information

Department of Chemistry, Pennsylvania State University, 104 Chemistry Building, University Park, Pennsylvania 16802, USA.

Erratum in

  • Biochemistry. 2005 Sep 13;44(36):12264.
  • Biochemistry. 2005 Mar 22;44(11):4600.


The T4 helicase-loading protein (gp59) has been proposed to coordinate leading- and lagging-strand DNA synthesis by blocking leading-strand synthesis during the primosome assembly. In this work, we unambiguously demonstrate through a series of biochemical and biophysical experiments, including single-molecule fluorescence microscopy, that the inhibition of leading-strand holoenzyme progression by gp59 is the result of a complex formed between gp59 and leading-strand polymerase (gp43) on DNA that is instrumental in preventing premature replication during the assembly of the T4 replisome. We find that both the polymerization and 3' --> 5' exonuclease activities of gp43 are totally inhibited within this complex. Chemical cross-linking of the complex followed by tryptic digestion and peptide identification through matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) mass spectrometry identified Cys169 of gp43 and Cys215 of gp59 as residues in a region of a protein-protein contact. With the available crystal structures for both gp43 and gp59, a model of the complex was constructed based on shape complementarity, revealing that parts of the C-terminal domain from gp59 insert into the interface created by the thumb and exonuclease domains of gp43. This insertion effectively locks the polymerase into a conformation where switching between the pol and editing modes is prevented. Thus, continued assembly of the replisome through addition of the primosome components and elements of the lagging-strand holoenzyme can occur without leading-strand DNA replication.

[Indexed for MEDLINE]

Publication type, MeSH terms, Substances, Grant support

Publication type

MeSH terms


Grant support

Supplemental Content

Full text links

Icon for American Chemical Society
Loading ...
Support Center