TY - JOUR
T1 - Proteins tightly associated with the termini of replicative form DNA of Kilham rat virus, an autonomous parvovirus
AU - Wobbe, C. R.
AU - Mitra, S.
PY - 1985
Y1 - 1985
N2 - Revie et al. [Revie, D., Tseng, B.Y., Grafstrom, R.H. & Goulian, M. (1979) Proc. Natl. Acad. Sci. USA 76, 5539-5543] have proposed that the double-stranded replicative form (RF) DNA of the autonomous rodent parvovirus H-1 has protein of 60 kDa covalently bound at its 5' termini. We present evidence that the RF DNA of a similar rodent parvovirus, Kilham rat virus (KRV), also has covalently bound protein. NaDodSO4/polyacrylamide gel electrophoresis of purified, 125I-labeled RF DNA shows that proteins of 68-72, 66, 64, and 55 kDa copurify with the DNA during velocity and equilibrium sedimentation in the presence of detergents and 4 M guanidine HCl. Phenol extraction in the presence of 2-mercaptoethanol removes the 68- to 72-kDa proteins, but the 66-, 64-, and 55-kDa proteins remain tightly, but noncovalently, bound. The latter polypeptides also appear to associate with protease-treated RF DNA when mixed with uninfected cell extract. Following removal of these proteins by electrophoresis in NaDodSO4/agarose gels, two proteins (called RF TP-90 and RF TP-40), of about 90 and 40 kDa, become evident. These remain bound to the DNA and are released only after nuclease digestion of the DNA. These two proteins, apparently not of viral origin, are associated with terminal restriction fragments of the RF DNA and appear to be covalently bound to the 5' termini of both strands.
AB - Revie et al. [Revie, D., Tseng, B.Y., Grafstrom, R.H. & Goulian, M. (1979) Proc. Natl. Acad. Sci. USA 76, 5539-5543] have proposed that the double-stranded replicative form (RF) DNA of the autonomous rodent parvovirus H-1 has protein of 60 kDa covalently bound at its 5' termini. We present evidence that the RF DNA of a similar rodent parvovirus, Kilham rat virus (KRV), also has covalently bound protein. NaDodSO4/polyacrylamide gel electrophoresis of purified, 125I-labeled RF DNA shows that proteins of 68-72, 66, 64, and 55 kDa copurify with the DNA during velocity and equilibrium sedimentation in the presence of detergents and 4 M guanidine HCl. Phenol extraction in the presence of 2-mercaptoethanol removes the 68- to 72-kDa proteins, but the 66-, 64-, and 55-kDa proteins remain tightly, but noncovalently, bound. The latter polypeptides also appear to associate with protease-treated RF DNA when mixed with uninfected cell extract. Following removal of these proteins by electrophoresis in NaDodSO4/agarose gels, two proteins (called RF TP-90 and RF TP-40), of about 90 and 40 kDa, become evident. These remain bound to the DNA and are released only after nuclease digestion of the DNA. These two proteins, apparently not of viral origin, are associated with terminal restriction fragments of the RF DNA and appear to be covalently bound to the 5' termini of both strands.
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U2 - 10.1073/pnas.82.24.8335
DO - 10.1073/pnas.82.24.8335
M3 - Article
C2 - 3866226
AN - SCOPUS:0022330612
SN - 0027-8424
VL - 82
SP - 8335
EP - 8339
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 24
ER -