Bacteriophage T4 RNA ligase: preparation of a physically homogeneous, nuclease-free enzyme from hyperproducing infected cells.

Abstract:

:Infection of Escherichia coli by a bacteriophage T4 regA, gene 44 double mutant leads to about a 7-fold increase in the amount of RNA ligase obtained after infection by wild-type phage. Using cells infected by the double mutant, RNA ligase was purified to homogeneity with a 20% yield. Unlike previous preparations of this enzyme, the ligase is free of contaminating nuclease and is therefore suitable for intermolecular ligation of DNA substrates. In the course of these studies it was discovered that adenylalation of the enzyme--a step in the reaction pathway--markedly decreased the electrophoretic mobility of RNA ligase through polyacrylamide gels containing sodium dodecyl sulfate. This behavior allows identification of RNA ligase among a mixture of proteins and was used to demonstrate that virtually all of the purified protein is enzymatically active.

journal_name

Nucleic Acids Res

journal_title

Nucleic acids research

authors

Higgins NP,Geballe AP,Snopek TJ,Sugino A,Cozzarelli NR

doi

10.1093/nar/4.9.3175

subject

Has Abstract

pub_date

1977-09-01 00:00:00

pages

3175-86

issue

9

eissn

0305-1048

issn

1362-4962

journal_volume

4

pub_type

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