Agarose gel electrophoresis reveals structural fluidity of a phage T3 DNA packaging intermediate

Philip Serwer, Elena T. Wright

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

We find a new aspect of DNA packaging-associated structural fluidity for phage T3 capsids. The procedure is (i) glutaraldehyde cross-linking of in vivo DNA packaging intermediates for the stabilization of structure and then (ii) determining effective radius by two-dimensional agarose gel electrophoresis (2D-AGE). The intermediates are capsids with incompletely packaged DNA (ipDNA) and without an external DNA segment; these intermediates are called ipDNA-capsids. We initially increase the production of ipDNA-capsids by raising NaCl concentration during in vivo DNA packaging. By 2D-AGE, we find a new state of contracted shell for some particles of one previously identified ipDNA-capsid. The contracted shell-state is found when the ipDNA length/mature DNA length (F) is above 0.17, but not at lower F. Some contracted-shell ipDNA-capsids have the phage tail; others do not. The contracted-shell ipDNA-capsids are explained by premature DNA maturation cleavage that makes accessible a contracted-shell intermediate of a cycle of the T3 DNA packaging motor. The analysis of ipDNA-capsids, rather than intermediates with uncleaved DNA, provides a simplifying strategy for a complete biochemical analysis of in vivo DNA packaging.

Original languageEnglish (US)
Pages (from-to)352-365
Number of pages14
JournalELECTROPHORESIS
Volume33
Issue number2
DOIs
StatePublished - Jan 2012

Keywords

  • Average electrical surface charge density
  • Bacteriophage DNA packaging motor
  • Chemical cross-linking
  • Effective particle radius
  • Two-dimensional electrophoresis

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry

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