Structural and functional characterization of the human DNA repair helicase XPD by comparative molecular modeling and site-directed mutagenesis of the bacterial repair protein UvrB.
A molecular model for the human nucleotide excision repair protein, XPD, was developed based on the structural and functional relationship of the protein with a bacterial nucleotide excision repair (NER) protein, UvrB. Whereas XPD does not share significant sequence identity with UvrB, the proteins share seven highly conserved helicase motifs that define a common protein structural template. They also have similar functional roles in their ATPase activity and the ability to unwind DNA and verify damaged strands in the process of NER. The validity of using the crystal structure of UvrB as a template for the development of an XPD model was tested by mimicking human disease-causing mutations (XPD: R112H, D234N, R601L) in UvrB (E110R, D338N, R506A) and by mutating two highly conserved residues (XPD, His-237 and Asp-609; UvrB, H341A and D510A). The XPD structural model can be employed in understanding the molecular mechanism of XPD human disease causing mutations. The value of this XPD model demonstrates the generalized approach for the prediction of the structure of a mammalian protein based on the crystal structure of a structurally and functionally related bacterial protein sharing extremely low sequence identity (<15%).
Bienstock
Rachelle J
RJ
Scientific Computing Laboratory, NIEHS, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
Skorvaga
Milan
M
Mandavilli
Bhaskar S
BS
Van Houten
Bennett
B
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000032:analysis* / Q000737:chemistry; D004265:DNA Helicases / Q000032:analysis* / Q000737:chemistry; D004260:DNA Repair; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D016415:Sequence Alignment; D014157:Transcription Factors; D051759:Xeroderma Pigmentosum Group D Protein
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D011506:Proteins; D014157:Transcription Factors; C058093:UvrB protein, E coli; D004265:DNA Helicases; D051759:Xeroderma Pigmentosum Group D Protein; C497292:ERCC2 protein, human