Preliminary X-ray crystallographic and NMR studies on the exonuclease domain of the epsilon subunit of Escherichia coli DNA polymerase III.
The structured core of the N-terminal 3'-5' exonuclease domain of epsilon, the proofreading subunit of Escherichia coli DNA polymerase III, was defined by multidimensional NMR experiments with uniformly (15)N-labeled protein: it comprises residues between Ile-4 and Gln-181. A 185-residue fragment, termed epsilon(1-185), was crystallized by the hanging drop vapor diffusion method in the presence of thymidine-5'-monophosphate, a product inhibitor, and Mn(2+) at pH 5.8. The crystals are tetragonal, with typical dimensions 0.2 mm x 0.2 mm x 1.0 mm, grow over about 2 weeks at 4 degrees C, and diffract X-rays to 2.0 A. The space group was determined to be P4(n)2(1)2 (n = 0, 1, 2, 3), with unit cell dimensions a = 60.8 A, c = 111.4 A.
Hamdan
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S
Research School of Chemistry, Australian National University, Canberra, ACT, 0200, Australia.
Brown
S E
SE
Thompson
P R
PR
Yang
J Y
JY
Carr
P D
PD
Ollis
D L
DL
Otting
G
G
Dixon
N E
NE
D020134:Catalytic Domain; D002918:Chymotrypsin / Q000378:metabolism; D003460:Crystallization; D018360:Crystallography, X-Ray; D004258:DNA Polymerase III / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology*; D005090:Exodeoxyribonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D016296:Mutagenesis; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D016415:Sequence Alignment; D013938:Thymidine Monophosphate / Q000378:metabolism
D010446:Peptide Fragments; D021122:Protein Subunits; D013938:Thymidine Monophosphate; D008345:Manganese; D004258:DNA Polymerase III; D005090:Exodeoxyribonucleases; D002918:Chymotrypsin