HUMAN HSP70 AND HSP40 CHAPERONE PROTEINS FACILITATE HUMAN PAPILLOMAVIRUS-11 E1 PROTEIN-BINDING TO THE ORIGIN AND STIMULATE CELL-FREE DNA-REPLICATION

Human papillomavirus replication initiator, the E1 helicase, binds wea kly to the origin of DNA replication. Purified human chaperone protein s Hsp70 and Hsp40 (HDJ-1 and HDJ-2) independently and additively enhan ced E1 binding to the origin. The interaction between E1 and Hsp70 was transient and required ATP hydrolysis, whereas Hsp40 bound to E1 dire ctly and remained in the complex. A peptide of 20 residues spanning th e HPD loop and helix II of the J domain of YDJ-1 also stimulated E1 bi nding to the origin, alone or in combination with Hsp70 or Hsp40. A mu tated peptide (H34Q) had a reduced activity, while an adjacent or an o verlapping peptide had no effect. Neither Hsp70 nor the J peptide alte red the E1/DNA ratio in the complex. Electron microscopy showed that E 1 mainly bound to DNA as a hexamer. In the presence of Hsp40, E1 prima rily bound to DNA as a dihexamer. Preincubation of chaperones with vir al E1 and template shortened the lag time and increased replication in a cell-free system. Since two helicases are essential for bidirection al replication of human papillomavirus DNA, these results demonstrate that, as in prokaryotes, chaperones play an important role in the asse mbly of preinitiation complexes on the origin.