Baynes J W, Wold F
J Biol Chem. 1976 Oct 10;251(19):6016-24.
The circulating half-lives of the four isozymes of bovine pancreatic ribonuclease (RNases A, B, C, and D) have been determined in normal and in nephrectomized rats. The isozymes differ only in their glycosyl content. While A contains no sugars, B has a simple oligosaccharide (GlcNAc2 Man4-5),and C and D each have a complex oligosaccharide (GlcNAc4 Man 2-3 Gal2 Fuc NeuAc2, and GlcNAc4 Man3 Gal2 Fuc NeuAc4, respectively) attached to Asn-34 of the polypeptide chain. All four isozymes were cleared rapidly in normal rats (t 1/2 = 2 to 3 min), as expected on the basis of the established role of the kidneys in removing low molecular weight proteins from circulation. In nephrectomized rats, however, a much slower clearance was observed, thus permitting the evaluation of the role of the carbohydrate chains in the catabolism of the isozymes. The clearance curves can be analyzed in terms of two processes, a rapid initial one, shown to represent the equilibration of the injected enzyme into extravascular space, and a second one which is interpreted as the catabolic clearance of the enzyme. The haf-life of the RNase isozymes was calculated from this second process and found to be in the range 528 to 577 min for RNase A, 15 min for RNase B, 681 to 862 min for RNase C, and 839 to 941 min for RNase D. The rapidly cleared RNase B was treated with alpha-mannosidase to remove 3 of the 4 mannosyl residues, leaving only a trisaccharide (GlcNAc2-betaMan) attached to the protein. The half-life of this RNase B derivatives was found to be in the range 616 to 733 min. From these results it is concluded (a) that the addition of complex oligosaccharides to a protein does not have any significant direct effect on its circulating half-life (RNases C and D compared to RNase A), and (b) that in the rat there exists a mechanism for clearing glycoproteins based on specific recognition of exposed alpha-mannosyl residues (RNase B compared to the other isozymes and to alpha-mannosidase-treated RNase B).
已测定牛胰核糖核酸酶的四种同工酶（核糖核酸酶A、B、C和D）在正常大鼠和肾切除大鼠体内的循环半衰期。这些同工酶仅糖基含量不同。A不含糖，B有一个简单寡糖（GlcNAc2Man4 - 5），C和D分别有一个复杂寡糖（分别为GlcNAc4Man2 - 3Gal2FucNeuAc2和GlcNAc4Man3Gal2FucNeuAc4）连接在多肽链的Asn - 34上。正如基于肾脏在从循环中清除低分子量蛋白质方面的既定作用所预期的那样，所有四种同工酶在正常大鼠体内清除迅速（t1/2 = 2至3分钟）。然而，在肾切除大鼠中，观察到清除速度慢得多，从而可以评估碳水化合物链在同工酶分解代谢中的作用。清除曲线可以根据两个过程进行分析，一个是快速的初始过程，已证明代表注入的酶在血管外空间达到平衡，另一个过程被解释为酶的分解代谢清除。核糖核酸酶同工酶的半衰期根据第二个过程计算得出，发现核糖核酸酶A为528至577分钟，核糖核酸酶B为15分钟，核糖核酸酶C为681至862分钟，核糖核酸酶D为839至941分钟。对快速清除的核糖核酸酶B用α - 甘露糖苷酶处理，去除4个甘露糖基残基中的3个，仅留下一个连接在蛋白质上的三糖（GlcNAc2 - βMan）。发现这种核糖核酸酶B衍生物的半衰期在616至733分钟范围内。从这些结果得出结论：（a）向蛋白质添加复杂寡糖对其循环半衰期没有任何显著直接影响（将核糖核酸酶C和D与核糖核酸酶A比较），（b）在大鼠中存在一种基于对暴露的α - 甘露糖基残基的特异性识别来清除糖蛋白的机制（将核糖核酸酶B与其他同工酶以及与α - 甘露糖苷酶处理的核糖核酸酶B比较）。
