DeoxyribonucleaseⅠ(DNaseⅠ) inhibitor of protein nature was isolated from bovine brain and was characterized to clarify the regulatory mechanisms of DNaseⅠactivity in vivo.
1. Two kinds of DNsae I inhibitor (peakⅠand peak Ⅱ) were isolated and purified to 15 fold for peakⅠand 26 fold for peakⅡ by centrifugation, ammonium sulfate fractionation, gel filtration and DEAE-cellulose chromatography. The molecular weights of peak I and peakⅡ inhibitors were estimated to be 160,000 and 42,000 dalton, respectively.
2. The maximum inhibitory activities of both peak I and peakⅡ inhibitors were observed at pH 7.5 and Mg^(2+) ion increased their inhibitory activities whereas Ca^(2+) and Mn^(2+) showed little effects on their inhibitory activities. The purified peaks I and Ⅱ inhibitors also revealed specific inhibitory activities to DNase I and did not exert any inhibitory effects on pancreatic RNase.
3. Peak I inhibitor protein was more stable than peakⅡ inhibitor protein to heat inactivation. Only 8% of peak I activiry disappeared by heat pretreatment at 45℃ for 10 minutes instead of 65% in case of peak Ⅱ.
4. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns for peakⅠ and peak Ⅱ inhibitor proteins revealed that the major protein peaks were located at the same position and molecular weight of the protein was estimated to be 42,000 dalton.
These results suggest the presence of a heat-stable DNase I inhibitor of protein nature in bovine brain which may consist of 4 monomeric units of actin, and well known DNase I inhibitor.
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