title: Structural and functional analysis of phi29 p16.7C dimerization mutants: identification of a novel aromatic cage dimerization motif.
abstract: Prokaryotic DNA replication is compartmentalized at the cellular membrane. The Bacillus subtilis phage varphi29-encoded membrane protein p16.7 is one of the few proteins known to be involved in the organization of prokaryotic membrane-associated DNA replication. The functional DNA binding domain of p16.7 is constituted by its C-terminal half, p16.7C, which forms high affinity dimers in solution and which can form higher order oligomers. Recently, the solution and crystal structures of p16.7C and the crystal structure of the p16.7C-DNA complex have been solved. Here, we have studied the p16.7C dimerization process and the structural and functional roles of p16.7 residues Trp-116 and Asn-120 and its last nine C-terminal amino acids, which form an extended tail. The results obtained show that transition of folded dimers into unfolded monomers occurs without stable intermediates and that both Trp-116 and the C-terminal tail are important for dimerization and functionality of p16.7C. Residue Trp-116 is involved in formation of a novel aromatic cage dimerization motif, which we call "Pro cage." Finally, whereas residue Asn-120 plays a minor role in p16.7C dimerization, we show that it is critical for both oligomerization and DNA binding, providing further evidence that DNA binding and oligomerization of p16.7C are coupled processes.
authors.affiliation: Instituto de Biología Molecular Eladio Viñuela (Consejo Superior de Investigaciones Científicas (CSIC)), Centro de Biología Molecular Severo Ochoa (CSIC-UAM (Universidad Autónoma de Madrid)), Universidad Autónoma, Canto Blanco, 28049 Madrid, Spain.
authors.forename: Daniel
authors.initials: D
authors.lastname: Muñoz-Espín
authors.forename: Miguel A
authors.initials: MA
authors.lastname: Fuertes
authors.forename: Mercedes
authors.initials: M
authors.lastname: Jiménez
authors.forename: Laurentino
authors.initials: L
authors.lastname: Villar
authors.forename: Carlos
authors.initials: C
authors.lastname: Alonso
authors.forename: Germán
authors.initials: G
authors.lastname: Rivas
authors.forename: Margarita
authors.initials: M
authors.lastname: Salas
authors.forename: Wilfried J J
authors.initials: WJ
authors.lastname: Meijer
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020816:Amino Acid Motifs / Q000235:genetics; D019943:Amino Acid Substitution; D017103:Bacillus Phages / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001412:Bacillus subtilis / Q000378:metabolism / Q000821:virology; D004261:DNA Replication / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D020125:Mutation, Missense; D020836:Protein Structure, Quaternary / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D013329:Structure-Activity Relationship; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D014764:Viral Proteins; D000251:Adenosine Triphosphatases; C439525:protein p16, bacteriophage phi29