The association and involvement of some members of the P1 protein family in a cell-free DNA replication of Xenopus eggs.

Two types of antibodies were prepared one directed against an oligopeptide specific to P1Cdc46, a mammalian homologue of yeast CDC46, and the other against an oligopeptide highly conserved in the P1 protein family. Immunoprecipitation with anti-P1Cdc46 antibody revealed that some members of the P1 protein family were coprecipitated with P1Cdc46 in the soluble fraction of Xenopus S phase extracts. Immunoblot analysis showed that all of the coprecipitated proteins reacted with the antibody against an oligopeptide, designated as a DEAD box motif, a highly conserved sequence in the P1 protein family. The immunodepleted extracts with anti-P1Cdc46 antibody-bound beads showed much lower activity of DNA replication than the mock-treated extracts. Recovery of replication was achieved by supplementing depleted extracts with the proteins eluted from anti-P1Cdc46 antibody-bound beads. These findings suggest that the proteins contained in the P1 protein family were associated in the extracts and that the multiprotein complex of the family plays an essential role in a cell-free DNA replication of Xenopus eggs.