A docking model based on mass spectrometric and biochemical data describes phage packaging motor incorporation.
The molecular mechanism of scaffolding protein-mediated incorporation of one and only one DNA packaging motor/connector dodecamer at a unique vertex during lambdoid phage assembly has remained elusive because of the lack of structural information on how the connector and scaffolding proteins interact. We assembled and characterized a phi29 connector-scaffolding complex, which can be incorporated into procapsids during in vitro assembly. Native mass spectrometry revealed that the connector binds at most 12 scaffolding molecules, likely organized as six dimers. A data-driven docking model, using input from chemical cross-linking and mutagenesis data, suggested an interaction between the scaffolding protein and the exterior of the wide domain of the connector dodecamer. The connector binding region of the scaffolding protein lies upstream of the capsid binding region located at the C terminus. This arrangement allows the C terminus of scaffolding protein within the complex to both recruit capsid subunits and mediate the incorporation of the single connector vertex.
Fu
Chi-yu
CY
Deptartment of Microbiology, University of Alabama, Birmingham, Alabama 35294-2170, USA.
Uetrecht
Charlotte
C
Kang
Sebyung
S
Morais
Marc C
MC
Heck
Albert J R
AJ
Walter
Mark R
MR
Prevelige
Peter E
PE
D000595:Amino Acid Sequence; D017103:Bacillus Phages / Q000502:physiology*; D001665:Binding Sites; D002213:Capsid / Q000737:chemistry; D036022:Capsid Proteins / Q000737:chemistry; D003432:Cross-Linking Reagents; D018360:Crystallography, X-Ray; D013058:Mass Spectrometry; D008958:Models, Molecular; D020409:Molecular Motor Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D017433:Protein Structure, Secondary; D015203:Reproducibility of Results; D019065:Virus Assembly / Q000502:physiology*
D036022:Capsid Proteins; D003432:Cross-Linking Reagents; D020409:Molecular Motor Proteins; D010455:Peptides