Structural analysis of DNA cleaved in vivo by bacteriophage T4 terminase.
Phage terminases are protein complexes that cleave concatemeric phage DNA and generate termini of the packaged DNA molecule. In phage T4, the DNA packaging proteins gp16 and gp17 are supposed to function as terminases. The recombinant T4 terminase proteins, upon expression in vivo from strong promoters, cleaved plasmid DNA in a sequence-independent manner. Resolution of the cleaved DNA by agarose-gel electrophoresis showed a smear throughout the lane including a fraction that was retained in the well [Bhattacharyya and Rao, Virology 196 (1993) 34-44]. The appearance of a smear in the high-M(r) region could not be explained solely on the basis of a simple random-cutting mechanism. Various hypotheses were tested to elucidate the structure of the high-M(r) DNA. The data show that the high-M(r) DNA did not arise either by attachment of protein(s) to DNA, or by covalent linkage of cleaved DNA molecules by a recombinational mechanism. It appears that the high-M(r) DNA arose as a result of non-covalent linkage of plasmid DNA through single strands. A working model for the action of T4 terminase is presented.
Bhattacharyya
S P
SP
Department of Biology, Catholic University of America, Washington, DC 20064.
Rao
V B
VB
D017122:Bacteriophage T4 / Q000201:enzymology* / Q000235:genetics; D004279:DNA, Viral / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D004706:Endodeoxyribonucleases / Q000737:chemistry*
D004279:DNA, Viral; D004706:Endodeoxyribonucleases; C030894:terminase