TY - JOUR
T1 - DNA gyrase
T2 - Affinity chromatography on novobiocin-sepharose and catalytic properties
AU - Staudenbauer, Walter L.
AU - Orr, Elisha
N1 - Funding Information:
ACKNOWLEDGEMENTS We thank Dr. Heinz Schuster for support and encouragement. The excellent technical assistance of Ursula Cinque is gratefully acknowledged. E. Orr was supported by a project grant from the Medical Research Council (U.K.) and a short term EMBO fellowship.
PY - 1981/8/11
Y1 - 1981/8/11
N2 - Novobiocin-Sepharose was prepared by coupling of novobiocin to Epoxy-activated Sepharose 6B and used as an affinity adsorbent. Four novobiocin-binding proteins were isolated from crude extracts of Escherichia coli with molecular weights of 105, 92, 85, and 40 kdal. The two larger proteins were identified as the A subunit (gyrA protein) and the B subunit (gyrB protein) of DNA gyrase (topoisomerase II). By this method the two gyrase components can be easily separated and purified in high yield. Although both proteins are involved in the ATP-dependent supercoiling of relaxed plasmid DNA, only the gyrB protein is required for catalyzing the cleavage of ATP. The gyrB protein ATPase activity is competitively inhibited by novobiocin and related coumarin antibiotics. ATP hydrolysis is unaffected by the addition of either gyrA protein or DNA but stimulated in the presence of both.
AB - Novobiocin-Sepharose was prepared by coupling of novobiocin to Epoxy-activated Sepharose 6B and used as an affinity adsorbent. Four novobiocin-binding proteins were isolated from crude extracts of Escherichia coli with molecular weights of 105, 92, 85, and 40 kdal. The two larger proteins were identified as the A subunit (gyrA protein) and the B subunit (gyrB protein) of DNA gyrase (topoisomerase II). By this method the two gyrase components can be easily separated and purified in high yield. Although both proteins are involved in the ATP-dependent supercoiling of relaxed plasmid DNA, only the gyrB protein is required for catalyzing the cleavage of ATP. The gyrB protein ATPase activity is competitively inhibited by novobiocin and related coumarin antibiotics. ATP hydrolysis is unaffected by the addition of either gyrA protein or DNA but stimulated in the presence of both.
UR - http://www.scopus.com/inward/record.url?scp=0019878168&partnerID=8YFLogxK
U2 - 10.1093/nar/9.15.3589
DO - 10.1093/nar/9.15.3589
M3 - Article
C2 - 6269086
AN - SCOPUS:0019878168
SN - 0305-1048
VL - 9
SP - 3589
EP - 3603
JO - Nucleic Acids Research
JF - Nucleic Acids Research
IS - 15
ER -