Purification and characterization of the Caenorhabditis elegans HCF protein and domains of human HCF.
The human cellular factor (HCF) is a multidomain protein that is implicated in processes of cell cycle progression, and it is recruited into a multicomponent assembly that triggers the expression of the herpes simplex virus genome. The amino-terminal domain of HCF has been proposed to form a "kelch" type beta-propeller fold, and the carboxy-terminal domain contains a repeat of a fibronectin-like motif. We describe the expression, purification, and characterization of the domains from the human HCF and of the full-length HCF from Caenorhabditis elegans. The purified recombinant C. elegans HCF can substitute for the human HCF in efficiently forming a multiprotein complex on a herpes simplex virus promoter element. As noted in earlier studies, a segment of human HCF encompassing the human kelch domain forms a stable complex on a viral promoter element. The purified fibronectin domain can also be recruited into this complex, but not into the stable complex formed with the minimal kelch domain. These results suggest that the fibronectin domain can interact with HCF in the transcriptional activating complex and that the association requires a region outside the putative beta-propeller.
Juang
Bi-Tzen
BT
Department of Biochemistry, University of Cambridge, UK.
Izeta
Ander
A
O'Hare
Peter
P
Luisi
Ben F
BF
D000818:Animals; D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics; D004330:Drosophila; D005353:Fibronectins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D017405:Herpes Simplex Virus Protein Vmw65 / Q000502:physiology; D051863:Host Cell Factor C1; D006801:Humans; D008840:Microfilament Proteins / Q000235:genetics / Q000502:physiology; D050811:Octamer Transcription Factor-1; D010446:Peptide Fragments / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D017510:Protein Folding; D017434:Protein Structure, Tertiary / Q000235:genetics; D021381:Protein Transport / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D012995:Solubility; D013347:Subcellular Fractions / Q000737:chemistry / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism
D029742:Caenorhabditis elegans Proteins; D004268:DNA-Binding Proteins; D005353:Fibronectins; C497692:HCFC1 protein, human; D017405:Herpes Simplex Virus Protein Vmw65; D051863:Host Cell Factor C1; C412617:KLHL1 protein, human; D008840:Microfilament Proteins; D050811:Octamer Transcription Factor-1; C494165:POU2F1 protein, human; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014157:Transcription Factors; C501364:fibronectin type III like peptide, human; C497691:hcf-1 protein, C elegans