Open AccessIn vitro assembly of the Bacillus subtilis bacteriophage phi 29.
Author(s) -
MaryAnn Bjornsti,
Bernard E. Reilly,
Dwight L. Anderson
Publication year - 1981
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.78.9.5861
Subject(s) - bacillus subtilis , bacteriophage , adenosine triphosphate , in vitro , dna , biology , biochemistry , in vivo , microbiology and biotechnology , gene , escherichia coli , bacteria , genetics
In vitro assembly of the Bacillus subtilis bacteriophage phi 29 that approaches the efficiency of assembly in vivo has been demonstrated. Proheads, DNA, and gene 16 product (gp16) were essential for DNA encapsidation, and the average yield in extracts was 180 phage per prohead donor cell. The in vitro maturation was very similar to in vivo assembly in terms of yield, intermediates, and abortive structures. More that 30% of the proheads in the extract were converted to phage, and about 20% of DNA--protein extracted from phage could be repackaged. In vitro assembly was blocked by the addition of DNase I, EDTA, pyrophosphatase, or the ATP analogues adenosine 5'-[alpha, beta-methylene]triphosphate and adenosine 5'-[beta, gamma-methylene]triphosphate. Less than 1% of the proheads isolated in sucrose gradients can accept DNA--protein in packaging in vitro.