Abstract
Abstract Extracts of wild type Escherichia coli contain a DNase which catalyzes the release of acid-soluble fragments from native DNA; this activity cannot be detected in rec B and C mutants. The properties of this enzyme have been studied with a fraction purified 200- to 600-fold. Only in the presence of ATP does the purified enzyme act on native DNA; with denatured DNA this ATP requirement is not absolute. The purified fraction requires Mg++ and is active over a broad pH range with an optimum of about 9.2. The exonuclease action of the purified enzyme fraction is due to two exonuclease activities. (a) An exonuclease, which we have called exonuclease V, acts on double-stranded DNA only in the presence of an activating nucleoside triphosphate, ATP and dATP being the most efficient. During DNA degradation, ATP is cleaved to ADP and Pi. The products formed during the degradation of native DNA are composed of 3'-hydroxyl- and 5'-phosphate-ended oligonucleotides, the average size of which decreases as the reaction proceeds. Exonuclease V attacks DNA from ends generated by double-stranded breakage and does not attack circular double-stranded DNA molecules with or without single-stranded breaks. Also associated with exonuclease V is an ATP-dependent cleavage of single-stranded DNA. (b) A 3'-exonuclease, acts on single-stranded DNA in the absence of ATP, and whose properties are distinguishable from those of exonuclease I, is responsible for the other exonuclease activity. The possible role of exonuclease V in DNA recombination is discussed.
Highlights
The exonuclease action of the purified enzyme fraction is due to two exonuclease activities. (a) A n exonuclease, which we have called exonuclease V, acts on double-stranded DNA only in the presence of an activating nucleoside triphosphate, ATP
The products formed during the degradation of native DNA are composed of 3’
The three other DNase activities present in the hydroxylapatite fraction are: (a) an exonucleolytic activity, acting on native DNA, which has an absolute dependence on,4TP
Summary
Strains and MediaThe genetic nomenclature used is according to the classification of Taylor and Trotter [20]. F- thr leu ser A str’ A end A 25 thi ret B 21, obtained by transduction of thy+A ret B 21 by bacteriophage Pl grown on AB 3069. Hfr 313 leu ilv thy R ret B 21 strr A, in MRW 36: a thy derivative of MRW 34 obtained without mutagenesis by a trimethoprim selection [21] ; MRW 51 F thr Zeu ser A str’ A thi end A 25 ret. C 22, obtained by transduction of thy+ A ret C 22 by bacteriophage PI grown on AB 3070 Hfr 313 leu ilv ret C 22 strr A, in MRW 36; AB 3069 and AB 3070 are gifts of Dr R. Media used for the growth of bacteria were the following: ML medium Bacto-tryptone, aud casamino acids were purchased from Difco Laboratories
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
