TY - JOUR
T1 - Protein-DNA cross-linking demonstrates stepwise ATP-dependent assembly of T4 DNA polymerase and its accessory proteins on the primer-template
AU - Capson, Todd L.
AU - Benkovic, Stephen J.
AU - Nossal, Nancy G.
N1 - Funding Information:
We thank Deborah Hinton, Howard Nash, and Kiyoshi Mizuuchi for helpful discussions and Linda Hoffman for typing the manuscript. This work was supported in part by grants from the National Institute of General Medical Sciences (GM 13306) and the National Foundation for Cancer Research to S. J. B. and by an NIH postdoctoral fellowship (GM 12011) to T. L. C.
PY - 1991/4/19
Y1 - 1991/4/19
N2 - T4 DNA polymerase, the 44 62 and 45 polymerase accessory proteins, and 32 single-stranded DNA-binding protein catalyze ATP-dependent DNA synthesis. Using DNA primers with cross-linkable residues at specific positions, we obtained structural data that reveal how these proteins assemble on the primer-template. With the nonhydrolyzable ATP analog ATPγS, assembly of the 44 62 and 45 proteins on the primer requires 32 protein but not polymerase. ATP hydrolysis changes the position and intensity of cross-linking to each of the accessory proteins and allows cross-linking of polymerase. Our data indicate that the initial binding of the three accessory proteins and ATP to a 32 protein-covered primer-template is followed by ATP hydrolysis, binding of polymerase, and movement of the accessory proteins to yield a complex capable of processive DNA synthesis.
AB - T4 DNA polymerase, the 44 62 and 45 polymerase accessory proteins, and 32 single-stranded DNA-binding protein catalyze ATP-dependent DNA synthesis. Using DNA primers with cross-linkable residues at specific positions, we obtained structural data that reveal how these proteins assemble on the primer-template. With the nonhydrolyzable ATP analog ATPγS, assembly of the 44 62 and 45 proteins on the primer requires 32 protein but not polymerase. ATP hydrolysis changes the position and intensity of cross-linking to each of the accessory proteins and allows cross-linking of polymerase. Our data indicate that the initial binding of the three accessory proteins and ATP to a 32 protein-covered primer-template is followed by ATP hydrolysis, binding of polymerase, and movement of the accessory proteins to yield a complex capable of processive DNA synthesis.
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U2 - 10.1016/0092-8674(91)90159-V
DO - 10.1016/0092-8674(91)90159-V
M3 - Article
C2 - 1849797
AN - SCOPUS:0025800987
SN - 0092-8674
VL - 65
SP - 249
EP - 258
JO - Cell
JF - Cell
IS - 2
ER -