Porcine purple acid phosphatase: heterologous expression, characterization, and proteolytic analysis

详细信息    查看全文

• 作者：Naseri ; Joseph Itor ; Truong ; Ngoc Thanh ; H??rentrup ; Jens ; Kuballa ; Petric ; Vogel ; Andreas ; et. al.
• 关键词：Uteroferrin ; Purple acid phosphatase ; Porcine iron transport ; Proteolysis ; Metalloenzyme ; Baculovirus
• 刊名：Archives of Biochemistry and Biophysics
• 出版年：2004
• 出版时间：December 1, 2004
• 年：2004
• 卷：432
• 期：1
• 页码：25-36
• 全文大小：652 K

文摘

Uteroferrin is an iron-binding glycoprotein, which is abundantly synthesized in porcine uterine glandular endometrium and believed to be involved in maternal/fetal iron transport. In the present study, uteroferrin has been cloned and functionally expressed using baculovirus-infected insect host cells Spodoptera frugiperda. The work also addresses the possible role of proteolytic cleavage to facilitate the release of uteroferrin-bound iron. The enzyme secreted in culture medium exhibits a molecular mass and catalytic properties similar to native porcine uteroferrin. The specific activity was estimated at 233U/mg using p-nitrophenyl phosphate as substrate. Partial cleavage of the enzyme with trypsin resulted in a 1.7-fold enhancement in specific activity and a two-subunit polypeptide as observed in preparations of most mammalian purple acid phosphatases. Digestion with the aspartic protease pepsin resulted in a 2.5-fold enzyme inactivation correlated with the appearance of low molecular weight polypeptide fragments and the release of enzyme-bound iron.