Half-life of N-acetylneuraminic acid in plasma membranes of rat liver and Morris hepatoma 7777.
Half-life of N-acetylneuraminic acid in plasma membranes of rat liver and Morris hepatoma 7777.
复制标题
大鼠肝脏和 Morris 肝癌 7777 质膜中 N-乙酰神经氨酸的半衰期。
作者：
E. Harms;W. Reutter
研究了大鼠肝脏和Morris肝癌7777细胞质膜和总蛋白中N -乙酰神经氨酸（NANA）的降解速率。Morris肝癌7777的质膜不能用已知的用于分离大鼠肝质膜的方法分离。因此，开发了一种改进的方法。作为质膜标记酶的5′-核苷酸酶的比活性相对于匀浆富集了12倍。N -乙酰-d-甘露糖胺是蛋白结合NANA的最合适的前体。当与使用具有可比比放射性的游离NANA相比时，掺入率在肝脏中高6倍，在肝癌中高20倍。N -乙酰基-d-甘露糖胺的乙酰基或己糖骨架中的14 C标记导致NANA半衰期的相同测量，导致NANA分子的均匀转换。NANA在肝细胞膜和肝癌细胞膜中的半衰期无差异。这同样适用于总肝脏和肝癌蛋白。然而，在两种组织中，质膜结合NANA的半衰期（肝脏，22.2至24小时;宿主肝脏，25.5小时; Morris肝癌7777; 23.7小时）短于总蛋白结合NANA的半衰期（肝脏，34小时;宿主肝脏，30.1小时; Morris肝癌7777，33.9小时）。这一发现表明，具有相当长的NANA半衰期的唾液酸蛋白存在于质膜以外的亚细胞结构中。在质膜中，NANA的半衰期比使用不可再利用的氨基酸前体时氨基酸的半衰期短得多。可以得出结论，NANA的平均周转比质膜中氨基酸的周转更快。质膜结合糖基转移酶的糖基化或质膜化合物的不同周转率被认为是导致质膜结合NANA半衰期较短的原因。
Summary The degradation rates of N -acetylneuraminic acid (NANA) were investigated in plasma membranes and in the total protein of rat liver and Morris hepatoma 7777. The plasma membranes of Morris hepatoma 7777 could not be isolated by known methods useful for isolation of rat liver plasma membranes. Therefore, a modified method was developed. The specific activity of 5′-nucleotidase as a plasma membrane marker enzyme was enriched twelvefold relative to homogenate. N -Acetyl-d-mannosamine is the most suitable precursor of protein-bound NANA. When compared to the use of free NANA of a comparable specific radioactivity, the rate of incorporation is 6 times higher in liver and 20 times higher in hepatoma. 14 C labeling in the acetyl group or in the hexose skeleton of N -acetyl-d-mannosamine resulted in identical measurement of the half-life of NANA, leading to a uniform turnover of the NANA molecule. There is no difference between the half-lives of NANA of the liver membranes and hepatoma membranes. The same applies to total liver and hepatoma protein. Yet, in both tissues, the half-life of plasma membrane-bound NANA is shorter (liver, 22.2 to 24 hr; host liver, 25.5 hr; Morris hepatoma 7777; 23.7 hr) than the half-life of total protein-bound NANA (liver, 34 hr; host liver, 30.1 hr; Morris hepatoma 7777, 33.9 hr). This finding indicates that sialoproteins with considerably longer half-lives of NANA exist in subcellular structures other than plasma membranes. In plasma membranes, the half-lives of NANA were much shorter than the half-lives of amino acids when a nonreutilizable amino acid precursor was used. It may be concluded that the mean turnover of NANA is more rapid than the turnover of amino acids in plasma membranes. Glycosylation by plasma membrane-bound glycosyltransferases or different turnover rates of plasma membrane compounds is regarded as responsible for the shorter half-life of plasma membrane-bound NANA.