Electrostatic interactions govern both nucleation and elongation during phage P22 procapsid assembly.
Icosahedral capsid assembly is an example of a reaction controlled solely by the interactions of the proteins involved. Bacteriophage P22 procapsids can be assembled in vitro by mixing coat and scaffolding proteins in a nucleation-limited reaction, where scaffolding protein directs the proper assembly of coat protein. Here, we investigated the effect of the buffer composition on the interactions necessary for capsid assembly. Different concentrations of various salts, chosen to follow the electroselectivity series for anions, were added to the assembly reaction. The concentration and type of salt was found to be crucial for proper nucleation of procapsids. Nucleation in low salt concentrations readily occurred but led to bowl-like partial procapsids, as visualized by negative stain electron microscopy. The edge of the partial capsids remained assembly-competent since coat protein addition triggered procapsid completion. The addition of salt to the partial capsids also caused procapsid completion. In addition, each salt affected both assembly rates and the extent of procapsid formation. We hypothesize that low salt conditions increase the coat protein:scaffolding protein affinity, causing excessive nuclei to form, which decreases coat protein levels leading to incomplete assembly.
Parent
Kristin N
KN
Department of Molecular and Cell Biology, Unit 3125, University of Connecticut, Storrs, CT 06269-3125, USA.
Doyle
Shannon M
SM
Anderson
Eric
E
Teschke
Carolyn M
CM
D017100:Bacteriophage P22 / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure*; D036022:Capsid Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000502:physiology*; D002942:Circular Dichroism; D004587:Electrophoresis, Agar Gel; D012965:Sodium Chloride / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D055672:Static Electricity; D014364:Tryptophan / Q000032:analysis; D014779:Virus Replication / Q000502:physiology*
D036022:Capsid Proteins; D012965:Sodium Chloride; D014364:Tryptophan