REDOR NMR characterization of DNA packaging in bacteriophage T4.
Bacteriophage T4 is a large-tailed Escherichia coli virus whose capsid is 120x86 nm. ATP-driven DNA packaging of the T4 capsid results in the loading of a 171-kb genome in less than 5 min during viral infection. We have isolated 50-mg quantities of uniform (15)N- and [epsilon-(15)N]lysine-labeled bacteriophage T4. We have also introduced (15)NH(4)(+) into filled, unlabeled capsids from synthetic medium by exchange. We have examined lyo- and cryoprotected lyophilized T4 using (15)N{(31)P} and (31)P{(15)N} rotational-echo double resonance. The results of these experiments have shown that (i) packaged DNA is in an unperturbed duplex B-form conformation; (ii) the DNA phosphate negative charge is balanced by lysyl amines (3.2%), polyamines (5.8%), and monovalent cations (40%); and (iii) 11% of lysyl amines, 40% of -NH(2) groups of polyamines, and 80% of monovalent cations within the lyophilized T4 capsid are involved in the DNA charge balance. The NMR evidence suggests that DNA enters the T4 capsid in a charge-unbalanced state. We propose that electrostatic interactions may provide free energy to supplement the nanomotor-driven T4 DNA packaging.
Yu
Tsyr-Yan
TY
Department of Chemistry, Washington University, St. Louis, MO 63130, USA.
Schaefer
Jacob
J
D000588:Amines / Q000737:chemistry; D017122:Bacteriophage T4 / Q000235:genetics*; D002412:Cations / Q000737:chemistry; D042003:DNA Packaging; D004279:DNA, Viral / Q000737:chemistry; D008958:Models, Molecular; D015394:Molecular Structure; D019906:Nuclear Magnetic Resonance, Biomolecular; D009690:Nucleic Acid Conformation; D010710:Phosphates / Q000737:chemistry; D011687:Purines / Q000737:chemistry; D011743:Pyrimidines / Q000737:chemistry
D000588:Amines; D002412:Cations; D004279:DNA, Viral; D010710:Phosphates; D011687:Purines; D011743:Pyrimidines