title: Length quantization of DNA partially expelled from heads of a bacteriophage T3 mutant.
abstract: DNA packaging of phages phi29, T3 and T7 sometimes produces incompletely packaged DNA with quantized lengths, based on gel electrophoretic band formation. We discover here a packaging ATPase-free, in vitro model for packaged DNA length quantization. We use directed evolution to isolate a five-site T3 point mutant that hyper-produces tail-free capsids with mature DNA (heads). Three tail gene mutations, but no head gene mutations, are present. A variable-length DNA segment leaks from some mutant heads, based on DNase I-protection assay and electron microscopy. The protected DNA segment has quantized lengths, based on restriction endonuclease analysis: six sharp bands of DNA missing 3.7-12.3% of the last end packaged. Native gel electrophoresis confirms quantized DNA expulsion and, after removal of external DNA, provides evidence that capsid radius is the quantization-ruler. Capsid-based DNA length quantization possibly evolved via selection for stalling that provides time for feedback control during DNA packaging and injection.
authors.affiliation: Department of Biochemistry, The University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, TX 78229-3900, USA. Electronic address: serwer@uthscsa.edu.
authors.forename: Philip
authors.initials: P
authors.lastname: Serwer
authors.affiliation: Department of Biochemistry, The University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, TX 78229-3900, USA.
authors.forename: Elena T
authors.initials: ET
authors.lastname: Wright
authors.affiliation: Markey Center for Structural Biology, Department of Biological Sciences, Purdue University, West Lafayette, IN 47907, USA.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Liu
authors.affiliation: Markey Center for Structural Biology, Department of Biological Sciences, Purdue University, West Lafayette, IN 47907, USA.
authors.forename: Wen
authors.initials: W
authors.lastname: Jiang
mesh_terms: D017903:Bacteriophage T3 / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure*; D004279:DNA, Viral / Q000378:metabolism* / Q000648:ultrastructure; D003850:Deoxyribonuclease I / Q000378:metabolism; D004586:Electrophoresis; D046529:Microscopy, Electron, Transmission; D009154:Mutation; D014771:Virion / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure*; D019065:Virus Assembly
chemical_list: D004279:DNA, Viral; D003850:Deoxyribonuclease I
keywords: Agarose gel electrophoresis, native; Bacteriophage assembly; Capsid structure; DNA packaging; Mutant phenotype; Ultracentrifugation