大鼠下颌下唾液腺黏液糖蛋白的酶促硫酸化作用。

Enzymatic sulfation of mucus glycoprotein in rat submandibular salivary gland.

Slomiany B L, Murty V L, Sarosiek J, Slomiany A

Dental Research Center, New Jersey Dental School, University of Medicine and Dentistry of New Jersey, Newark 07103-2425.

Int J Biochem. 1989;21(2):165-71. doi: 10.1016/0020-711x(89)90105-5.

Abstract

The enzymic activity which catalyzes transfer of sulfate ester group from 3'-phosphoadenosine-5'-phosphosulfate to mucus glycoprotein was found associated with Golgi-rich membrane fraction of rat submandibular salivary gland. 2. Optimum enzyme activity was obtained with 0.5% Triton X-100, 4 mM MgCl2 and 25 mM NaF at a pH of 6.8 using desulfated submandibular salivary mucus glycoprotein. The apparent Km of the enzyme for mucus glycoprotein was 11.1 mg/ml. 3. Alkaline borohydride reductive cleavage of the synthesized 35S-labeled glycoprotein led to the liberation of the label into reduced oligosaccharides. A 75.4% of the label was found incorporated in four oligosaccharides. These were identified in order of abundance as sulfated penta-, tri-, hepta- and nonsaccharides. 4. Based on the results of chemical and enzymatic analyses of the intact and desulfated compounds the pentasaccharide was characterized as SO3H----GlcNAc beta----Gal beta----GlcNAc(NeuAc alpha----)GalNAc-ol and the trisaccharide as SO3H----GlcNAc beta----Gal beta----GalNAc-ol.

摘要

发现催化硫酸酯基团从3'-磷酸腺苷-5'-磷酸硫酸转移至黏液糖蛋白的酶活性与大鼠下颌下唾液腺富含高尔基体的膜组分相关。2. 使用脱硫酸的下颌下唾液黏液糖蛋白，在pH为6.8时，加入0.5% Triton X-100、4 mM MgCl₂和25 mM NaF可获得最佳酶活性。该酶对黏液糖蛋白的表观Km为11.1 mg/ml。3. 合成的35S标记糖蛋白经碱性硼氢化物还原裂解后，标记物释放到还原的寡糖中。发现75.4%的标记物掺入四种寡糖中。按丰度顺序鉴定为硫酸化的五糖、三糖、七糖和非糖。4. 根据完整和脱硫酸化合物的化学及酶分析结果，五糖的特征为SO₃H----GlcNAcβ----Galβ----GlcNAc(NeuAcα----)GalNAc-ol，三糖的特征为SO₃H----GlcNAcβ----Galβ----GalNAc-ol。