THE XERODERMA-PIGMENTOSUM GROUP-B PROTEIN ERCC3 PRODUCED IN THE BACULOVIRUS SYSTEM EXHIBITS DNA HELICASE ACTIVITY

The XPB/ERCC3 gene corrects the nucleotide excision-repair defect in t he human hereditary disease xeroderma pigmentosum group B and encodes the largest subunit of the basal transcription factor BTF2/TFIIH. The primary sequence of the XPB/ERCC3 protein features the hallmarks of se ven helicase motifs found in many known and putative helicases or heli case-related proteins. Recently, the multiprotein BTF2/TFIIH complex h as been found to be associated with DNA helicase activity. To explore the properties and functions of XPB/ERCC3, we have used the baculoviru s/insect-cell expression system to produce recombinant protein. We rep ort here the construction and analysis of recombinant baculovirus expr essing XPB/ERCC3. The XPB/ERCC3 protein is synthesized at a relatively high level in baculovirus-infected insect cells. While the majority o f XPB/ERCC3 end up in the insoluble fraction of insect cell lysates, a minor fraction of recombinant protein is present in soluble form whic h can be purified under native conditions. We have found that a DNA he licase activity is associated with the purified XPB/ERCC3 protein, sug gesting that XPB/ERCC3 may function as a DNA helicase in local unwindi ng of DNA template both in the context of transcription and nucleotide excision repair.