Structural Insights to Insulin Receptor Ligand Interactions

摘要 胰岛素治疗极大地改善了糖尿病患者的健康，通常是每天一次。 长效胰岛素和正餐速效胰岛素。尽管取得了相当大的成功，但一些重要的 挑战依然存在，包括本提案中涉及的三个主要限制。首先，即使是最好的 临床上可用的速效胰岛素速度太慢，持续时间太长，无法严格控制血糖 在生理范围内，导致超出该范围的大量漂移和慢性高血糖 或急性低血糖并发症。其次，目前可用的胰岛素需要持续冷藏以避免 聚集，而不需要冷链输送的治疗性胰岛素将提供相当大的 优点，特别是用于长期胰岛素泵和在电力 不可靠。第三，因为胰岛素刺激两种信号，其中一种在治疗上有利于新陈代谢控制 (AKT途径)和一个与有丝分裂生长有关的治疗途径(ERK途径)，人们对 开发优先刺激Akt途径的类似物。这项提议需要一个生化和 基于结构的方法，以获得对每个问题的机械性洞察，包括冷冻-EM结构 多种方法辅助的受体-配体复合体的测定，包括细胞信号转导 以及小鼠的葡萄糖反应性研究。目的1重点研究锥形蜗牛蛇毒的两个人源化变体 胰岛素，它缺乏残留物，使天然胰岛素二聚体，并天生对皮下作用缓慢 注射，并已被设计为提供快速反应，作用持续时间短，和高效力。目标2 重点介绍使胰岛素抵抗聚集/纤颤的方法，包括对 令人惊讶的发现，其中一种人化的毒液胰岛素高度抵抗聚集。《目标3》探索 某些受体配基的显著特性，可以引发偏向的信号，强调Akt或 ERK途径，并为理解这些效应的结构基础提供了可能性。完成这些目标 将提供基本的机械洞察力，并为开发改进的疗法提供信息。

ABSTRACT Insulin treatment dramatically improves the health of people with diabetes, and is usually administered as a daily long-acting insulin and a prandial fast-acting insulin. Despite considerable success, a number of important challenges remain, including three major limitations that are addressed in this proposal. First, even the best clinically-available fast-acting insulins are too slow and last too long to provide tight control of serum glucose within the physiological range, resulting in substantial excursions outside of this range and chronic hyperglycemia or acute hypoglycemic complications. Second, currently available insulins require continual refrigeration to avoid aggregation, whereas therapeutic insulins that do not require cold-chain delivery would offer considerable advantages, especially for use in long-term insulin pumps and in circumstances where electrical power is unreliable. Third, because insulin stimulates two signals, one therapeutically advantageous for metabolic control (Akt pathway) and one therapeutically concerning for mitogenic growth (Erk pathway), there is interest in developing analogs that preferentially stimulate the Akt pathway. This proposal takes a biochemical and structure-based approach to gain mechanistic insight to each of these concerns, including cryo-EM structure determination of receptor-ligand complexes complemented by a variety of approaches, including cell signaling and mouse glucose-responsiveness studies. Aim 1 focusses on two humanized variants of cone snail venom insulins, which lack residues that make native insulin dimeric and inherently slow acting upon subcutaneous injection, and have been engineered to provide fast response, short duration of action, and high potency. Aim 2 focusses on approaches to render insulins resistant to aggregation/fibrillation, including following up on the surprising finding that one of the humanized venom insulins is highly resistant to aggregation. Aim 3 explores the remarkable property of some receptor ligands to elicit biased signaling that emphasizes either the Akt or the Erk pathways, and offers potential to understand the structural basis for these effects. Completion of these aims will provide fundamental mechanistic insights and inform efforts to develop improved therapeutics.