Single-Chain Insulins as Receptor Agonists
Single-Chain Insulins as Receptor Agonists
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DOI：
10.1210/me.2008-0349
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发表时间：
2009-05-01
影响因子：
--
通讯作者：
Arvan, Peter
中科院分区：
文献类型：
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作者：
Rajpal, Gautam;Liu, Ming;Arvan, Peter
单链胰岛素（sci）是一种单肽链，其中胰岛素b链与胰岛素a链通过未切割的连接肽连续连接。虽然胰岛素B链和胰岛素a链的直接连接产生的胰岛素受体结合很少，但生物学家一直对生物工程连接肽感兴趣，这种连接肽可以形成灵活的反向旋转，使胰岛素受体激活。在这篇报告中，我们研究了一系列cdna，旨在探索连接体长度、可切割性的重要性，以及某些位点依赖残基对重组SCIs对胰岛素受体的生物活性的影响。用（胰岛素原+胰岛素）特异性多克隆抗体，可以很容易地用RIA测定SCI浓度。尽管双碱基侧翼残基可能导致潜在的内蛋白水解易感性，但具有- gln - arg侧翼序列的连接体即使在分泌颗粒中也能抵抗裂解，从而确保单链行为。有效的SCIs与胰岛素受体具有良好的特异性结合。连接体在a链之前含有Arg残基的SCI具有最高的生物活性，尽管随着SCI连接体长度的增加，有效的受体相互作用受到抑制，接近胰岛素原。SCIs激活下游代谢信号，刺激脂肪细胞对葡萄糖的摄取，抑制肝细胞中糖异生酶的生物合成，仅对IGF-I受体具有有限的交叉反应性。从理论上讲，SCIs可能在胰岛素缺乏型糖尿病的免疫治疗或基因治疗中具有实用价值。（分子内分泌学23:679-688,2009）
Single-chain insulins (SCIs) are single polypeptide chains in which the insulin B-chain links contiguously with the insulin A-chain via an uncleaved connecting peptide. Although direct linkage of insulin B- and A-chains produces SCIs with little insulin receptor binding, biologists have been interested in bioengineering linker peptides that form a flexible reverse turn, allowing SCIs to activate insulin receptors. In this report, we have investigated a series of cDNAs intended to explore the significance of linker length, cleavability, and the impact of certain site-dependent residues for the bioactivity of recombinant SCIs on insulin receptors. SCI concentration is readily measured by RIA with a (proinsulin plus insulin)-specific polyclonal antibody. Although dibasic flanking residues may result in potential endoproteolytic susceptibility, a linker with -Gln-Arg-flanking sequences resisted cleavage even in secretory granules, ensuring single-chain behavior. Effective SCIs exhibit favorable and specific binding with insulin receptors. SCIs with linkers bearing an Arg residue immediately preceding the A-chain were most bioactive, although efficient receptor interaction was inhibited as SCI linker length increased, approaching that observed for proinsulin. SCIs activate downstream metabolic signaling, stimulating glucose uptake into adipocytes and suppressing gluconeogenic enzyme biosynthesis in hepatocytes, with only limited cross-reactivity on IGF-I receptors. SCIs might theoretically have utility either in immunotherapy or gene therapy in insulin-deficient diabetes. (Molecular Endocrinology 23: 679-688, 2009)