Structural insight into dynamic bypass of the major cisplatin-DNA adduct by Y-family polymerase Dpo4.
Y-family DNA polymerases bypass Pt-GG, the cisplatin-DNA double-base lesion, contributing to the cisplatin resistance in tumour cells. To reveal the mechanism, we determined three structures of the Y-family DNA polymerase, Dpo4, in complex with Pt-GG DNA. The crystallographic snapshots show three stages of lesion bypass: the nucleotide insertions opposite the 3'G (first insertion) and 5'G (second insertion) of Pt-GG, and the primer extension beyond the lesion site. We observed a dynamic process, in which the lesion was converted from an open and angular conformation at the first insertion to a depressed and nearly parallel conformation at the subsequent reaction stages to fit into the active site of Dpo4. The DNA translocation-coupled conformational change may account for additional inhibition on the second insertion reaction. The structures illustrate that Pt-GG disturbs the replicating base pair in the active site, which reduces the catalytic efficiency and fidelity. The in vivo relevance of Dpo4-mediated Pt-GG bypass was addressed by a dpo-4 knockout strain of Sulfolobus solfataricus, which exhibits enhanced sensitivity to cisplatin and proteomic alterations consistent with genomic stress.
Wong
Jimson H Y
JH
Department of Biochemistry, University of Western Ontario, London, Ontario, Canada.
Brown
Jessica A
JA
Suo
Zucai
Z
Blum
Paul
P
Nohmi
Takehiko
T
Ling
Hong
H
D000970:Antineoplastic Agents / Q000494:pharmacology; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020134:Catalytic Domain; D002470:Cell Survival; D002945:Cisplatin / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D018360:Crystallography, X-Ray; D018736:DNA Adducts / Q000737:chemistry / Q000378:metabolism*; D019641:DNA, Archaeal / Q000096:biosynthesis* / Q000737:chemistry; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D055786:Gene Knockout Techniques; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D020543:Proteome / Q000032:analysis; D048229:Sulfolobus solfataricus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D000970:Antineoplastic Agents; D019843:Archaeal Proteins; D018736:DNA Adducts; D019641:DNA, Archaeal; D020543:Proteome; C057270:cisplatin-DNA adduct; D004259:DNA-Directed DNA Polymerase; D002945:Cisplatin