Mutations in Sensor 1 and Walker B in the bovine papillomavirus E1 initiator protein mimic the nucleotide-bound state.
Viral replication initiator proteins are multifunctional proteins that utilize ATP binding and hydrolysis by their AAA+ modules for multiple functions in the replication of their viral genomes. These proteins are therefore of particular interest for understanding how AAA+ proteins carry out multiple ATP driven functions. We have performed a comprehensive mutational analysis of the residues involved in ATP binding and hydrolysis in the papillomavirus E1 initiator protein based on the recent structural data. Ten of the eleven residues that were targeted were defective for ATP hydrolysis, and seven of these were also defective for ATP binding. The three mutants that could still bind nucleotide represent the Walker B motif (D478 and D479) and Sensor 1 (N523), three residues that are in close proximity to each other and generally are considered to be involved in ATP hydrolysis. Surprisingly, however, two of these mutants, D478A and N523A, mimicked the nucleotide bound state and were capable of binding DNA in the absence of nucleotide. However, these mutants could not form the E1 double trimer in the absence of nucleotide, demonstrating that there are two qualitatively different consequences of ATP binding by E1, one that can be mimicked by D478A and N523A and one which cannot.
Liu
Xiaofei
X
Cold Spring Harbor Laboratory, P.O. Box 100, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.
Stenlund
Arne
A
D000255:Adenosine Triphosphate / Q000378:metabolism; D020816:Amino Acid Motifs; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D052688:Bovine papillomavirus 1 / Q000235:genetics* / Q000502:physiology*; D002417:Cattle; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics* / Q000502:physiology; D006868:Hydrolysis; D066298:In Vitro Techniques; D008958:Models, Molecular; D046912:Multiprotein Complexes; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics* / Q000502:physiology; D014779:Virus Replication
D004279:DNA, Viral; D004268:DNA-Binding Proteins; C069814:E1 protein, Bovine papillomavirus; C051550:E2 protein, Bovine papillomavirus; D046912:Multiprotein Complexes; D011994:Recombinant Proteins; D014764:Viral Proteins; D000255:Adenosine Triphosphate