Oxidative Folding of Peptides with Cystine‐Knot Architectures: Kinetic Studies and Optimization of Folding Conditions

Bioactive peptides often contain several disulfide bonds that provide the main contribution to conformational rigidity and structural, thermal, or biological stability. Among them, cystine‐knot peptides—commonly named “knottins”—make up a subclass with several thousand natural members. Hence, they are considered promising frameworks for peptide‐based pharmaceuticals. Although cystine‐knot peptides are available through chemical and recombinant synthetic routes, oxidative folding to afford the bioactive isomers still remains a crucial step. We therefore investigated the oxidative folding of ten protease‐inhibiting peptides from two knottin families, as well as that of an HIV entry inhibitor and of aprotinin, under two conventional sets of folding conditions and by a newly developed procedure. Kinetic studies identified folding conditions that resulted in correctly folded miniproteins with high rates of conversion even for highly hydrophobic and aggregation‐prone peptides in concentrated solutions.     

磁性亲合分离法纯化胰蛋白酶

采用壳聚糖包埋纳米钡铁氧体,经环氧氯丙烷交联制得磁性亲和载体,应用激光粒度分析仪和振动样品磁性测定仪对磁性亲和载体进行表征,动态激光散射分析结果表明,磁性粒子的平均直径为401纳米.磁性测量表明,超细磁粉具有较高的磁性.然后利用碳二酰亚胺活化法,将抑肽酶与此微粒共价结合得到磁性亲和吸附剂,应用于胰蛋白酶的亲和纯化.论文详细讨论胰蛋白酶亲和纯化条件,将磁性亲合分离法成功地应用于亲和纯化胰蛋白酶.     

抑肽酶对体外循环手术患者的影响

目的探讨体外循环手术患者应用抑肽酶的临床效果及安全性。方法以中国食品及药品监督局下令暂停抑肽酶在心脏外科临床使用的日期（2007年12月19日）为分界点,回顾性分析停药前后1年的患者临床资料。选择2006年12月19日至2007年12月18日行体外循环手术患者为抑肽酶组（n=360）,在术中均应用抑肽酶;2007年12月19日至2008年12月18日行体外循环手术患者为对照组（n=360）,在术中均未使用抑肽酶。对2组患者体外循环、主动脉阻断时间,术后出血量、输血量、呼吸机使用时间、术后24 h氧分压指数,二次开胸、术后肾功能不全、术后低心排出量综合征、延迟拔管、术后严重肺部感染等发生率及住院病死率进行比较。结果 2组患者手术种类、体外循环时间、主动脉阻断时间,肾功能不全、低心排出量综合征发生率,住院病死率的比较差异均无统计学意义（均P〉0.05）。抑肽酶组术后出血量、输血量、呼吸机使用时间,二次开胸、延迟拔管、严重肺部感染发生率均低于对照组（均P〈0.05）;术后24 h氧分压指数高于对照组（P〈0.05）。结论体外循环手术应用抑肽酶是安全有效的,可明显减少术后出血量及输血量,且对肺功能有一定的保护作用,并没有增加病死率及并发症发生率。所以,在中国是否应该停用抑肽酶还需要国内做一些大样本、多中心的临床研究来验证。     

琼脂糖凝胶提取纯化抑肽酶的研究

采用琼脂糖凝胶为载体,共价偶联牛胰蛋白酶,制成抑肽酶亲和吸附剂,将其用于亲和层析牛肺提取液,分离纯化高比活抑肽酶。通过实验研究,计算抑肽酶抑制比活单位(BAEE单位/mg胰蛋白酶)为79000,酶活性回收率76.63%。该工艺具有操作简便、收率高、产品质量好、污染低等优点,适于工业化,有一定的工业应用前景。     

Characterization and application of high magnetic property chitosan particles

High magnetic property chitosan particles were prepared from chitosan coated on magnetic powder, glutaraldehyde modified, and then epichlorohydrin crosslinked. The magnetic properties, the surface, and the morphology of the magnetic particles were characterized. An optimum condition of immobilization of aprotinin on magnetic particles was obtained, then these particles were used for affinity purification of trypsin, and the condition of affinity purification was discussed. The proposed method was successfully applied to the purification of trypsin. © 2001 John Wiley & Sons, Inc. J Appl Polym Sci 81: 1175–1181, 2001     

Preparation of highly magnetic chitosan particles and their use for affinity purification of enzymes

A novel protocol for preparation of highly magnetic chitosan particles, with a coercive force as high as 3500 Oe, has been developed. The surface of the particles was functionalized with aldehyde groups to facilitate the attachment of affinity ligands. The optimum conditions for the preparation of highly magnetic chitosan particles and immobilization of trypsin on magnetic particles were obtained; these particles were then used for affinity purification of aprotinin, and the conditions of affinity purification are discussed in detail. The proposed method was successfully applied to the affinity purification of aprotinin. Copyright © 2003 Society of Chemical Industry