Neutral Deoxyribonucleases of HeLa S3 Cells

Abstract

Extracts of HeLa [human cervical cancer cells] S3 cells were electrophoresed on polyacrylamide gels; gel slices were eluted and the elutes were assayed for DNase activities against native and denatured DNA substrates in the presence of MgCl2 or Na2EDTA. Aliquots of each elutate were assayed for their ability to nick the circular supercoiled PM2 phage DNA to distinguish endonucleases from exonucleases. Peaks of endonuclease activities were characterized as forming 3''-phospho-oligonucleotides or 5''-phospho-oligonucleotides by the use of oligonucleotides produced by these enzymes as substrates for the 5''-phosphate-specific snake venom exonuclease. The total activity of DNases in gel eluates was much higher than that in cell extract applied to the gel, indicating the presence of inhibitors in the cell extract.