Purification and Characterzation of a Restriction Endonuclease from Pseudomonas syringae pv.phaselicola

Pseudomonas syringe pv. phaseolicola로 부터 제한효소의 분리정제 및 특성

  • Bae, Moo (Department of Biological Science, Ewha Woman's University) ;
  • Lee, Eun-Young (Department of Biological Science, Ewha Woman's University)
  • 배무 (이화여자대학교 생물과학과) ;
  • 이은영 (이화여자대학교 생물과학과)
  • Published : 1994.10.01

Abstract

A restriction endonuclease, PsyI, has been isolated from Pseudomonas syringae pv. pha- seolicola, and its catalytic properties have been studied. This enzyme was purified through strepto- mycin sulfate and ammonium sulfate fractionation, phosphocellulose Pll, DEAE-cellulose, hydroxy- apatite and Sephadex G-100 column chromatography. It's molecular weight was about 50,000 dalton as determined by 7.5% polyacrylamide gel electrophoresis containing 0.1% SDS. In catalytic proper- ties, PsyI shows stable at wide ranges of pH between 7.0 and 10.0, of temperature between 30$\circ$C and 37$\circ$C, and its thermal stability is between 25$\circ$C, and 45$\circ$C, at the presence Of 10 mM MgCl$_{2}$-PsyI essentially require Na salt for enzyme reaction, is rather inhibited in the high Na salt concent- ration. The presence of 2-mercaptoethanol is absolutely required for the enzyme activity. This endonuclease, PsyI was determined to be an isoschizomer of SalI from the results of the restriction mapping and DNA sequencing.

Keywords

Restriction endonuclease SalI;Pseudomonas syringae PsyI

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