Domena J D, Mosbaugh D W
Biochemistry. 1985 Dec 3;24(25):7320-8. doi: 10.1021/bi00346a045.
Rat liver uracil-DNA glycosylase has been purified from nuclear extracts over 3000-fold to apparent homogeneity as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme is a monomeric protein with a polypeptide molecular weight of approximately 35 000. It has a native molecular weight of 33 000 as determined by gel filtration chromatography and a sedimentation coefficient of 2.6 S in glycerol gradients. The nuclear enzyme has an alkaline pH optimum and a pI value of 9.3. Nuclear uracil-DNA glycosylase catalyzes the release of free uracil from both single-stranded and double-stranded DNA with the former being the preferred substrate. The enzyme is unable to recognize dUTP, dUMP, or poly(dA-dT) containing a 3'-terminal uracil residue as a substrate. However, internalization of terminal uracil residues by limited chain elongation produced a substrate for the glycosylase. Another species of uracil-DNA glycosylase has been partially purified from mitochondria. This activity differs from the nuclear enzyme in that it has (i) distinctive chromatographic properties, (ii) a lower native molecular weight of 20 000 as determined by molecular sieving, (iii) a distinct NaCl inhibition profile, and (iv) a longer half-life during thermal denaturation.
大鼠肝脏尿嘧啶 - DNA糖基化酶已从核提取物中纯化出来，经十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳测定，纯化倍数超过3000倍，达到表观均一性。该酶是一种单体蛋白，多肽分子量约为35000。通过凝胶过滤色谱法测定其天然分子量为33000，在甘油梯度中的沉降系数为2.6 S。核酶的最适pH为碱性，pI值为9.3。核尿嘧啶 - DNA糖基化酶催化从单链和双链DNA中释放游离尿嘧啶，前者是首选底物。该酶无法将含有3'-末端尿嘧啶残基的dUTP、dUMP或聚（dA - dT）识别为底物。然而，通过有限的链延伸使末端尿嘧啶残基内化后产生了一种糖基化酶的底物。另一种尿嘧啶 - DNA糖基化酶已从线粒体中部分纯化出来。这种活性与核酶不同之处在于它具有：（i）独特的色谱特性；（ii）通过分子筛测定的较低天然分子量20000；（iii）独特的NaCl抑制曲线；（iv）热变性期间更长的半衰期。
