PURIFICATION OF OVEREXPRESSED GAM GENE PROTEIN FROM BACTERIOPHAGE MU BY DENATURATION RENATURATION TECHNIQUES AND A STUDY OF ITS DNA-BINDING PROPERTIES

Recombinant Mu gam gene protein (Mu GAM) synthesized in Escherichia coli accumulates in the form of insoluble inclusion bodies which, after cell lysis and low-speed centrifugation, can be recovered in the pellet fraction. This property was utilized in a purification procedure for Mu GAM based on guanidine hydrochloride denaturation-renaturation followed by a single DEAE-cellulose chromatographic step. The purified Mu GAM was shown by nitrocellulose-filter-binding experiments to bind with high affinity to linear double-stranded DNA and more weakly to supercoiled and single stranded forms. Mu GAM protects linear DNA from degradation by a variety of exonucleases, but only weakly inhibits endonuclease activity. The results are in accord with a model of Mu GAM conferring protection from exonuclease activity by binding to the ends of DNA.