Efficient strand transfer by the RadA recombinase from the hyperthermophilic archaeon Desulfurococcus amylolyticus.
The radA gene predicted to be responsible for homologous recombination in a hyperthermophilic archaeon, Desulfurococcus amylolyticus, was cloned, sequenced, and overexpressed in Escherichia coli cells. The deduced amino acid sequence of the gene product, RadA, was more similar to the human Rad51 protein (65% homology) than to the E. coli RecA protein (35%). A highly purified RadA protein was shown to exclusively catalyze single-stranded DNA-dependent ATP hydrolysis, which monitored presynaptic recombinational complex formation, at temperatures above 65 degrees C (catalytic rate constant of 1.2 to 2.5 min(-1) at 80 to 95 degrees C). The RadA protein alone efficiently promoted the strand exchange reaction at the range of temperatures from 80 to 90 degrees C, i.e., at temperatures approaching the melting point of DNA. It is noteworthy that both ATP hydrolysis and strand exchange are very efficient at temperatures optimal for host cell growth (90 to 92 degrees C).
Kil
Y V
YV
Division of Molecular and Radiation Biophysics, Petersburg Nuclear Physics Institute, Russian Academy of Sciences, Gatchina/St. Petersburg 188350, Russia.
Baitin
D M
DM
Masui
R
R
Bonch-Osmolovskaya
E A
EA
Kuramitsu
S
S
Lanzov
V A
VA
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D003001:Cloning, Molecular; D004254:DNA Nucleotidyltransferases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D019620:Desulfurococcaceae / Q000201:enzymology* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D051135:Rad51 Recombinase; D011938:Rec A Recombinases / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid
D019843:Archaeal Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C117252:RadA protein, archaeal; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D004254:DNA Nucleotidyltransferases; C495153:RAD51 protein, human; D051135:Rad51 Recombinase; D011938:Rec A Recombinases