Design and Functional Evaluation of Albumin Mutants for Improving Medicinal Efficacy

提高药效的白蛋白突变体的设计和功能评价

• 批准号: 10557245
• 负责人: OTAGIRI Masaki
• 金额: $ 6.85万
• 依托单位: KUMAMOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B).
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-10557245/
• 关键词: Albumin preparation; Site-directed mutagenesis; Molecular design; Artificial blood; Functional evaluation; Blood preparation; アルブミン

This investigation was undertaken to develop a safe and effective albumin using site-directed mutagenesis techniques. The results obtained in this study are as follow :1) Two single-mutants (K199A, K525A) were produced. The recombinant albumins were correctly folded, as evidenced by the fact that they showed similar patterns as the native albumin in the far- and near-UV CD spectrum as well as reactivity towards polyclonal anti-serum raised against native albumin.2) Thermal denaturation experiments by using GdnCl and DSC revealed no significant difference in conformational stability for native and mutant albumins.3) The ligand binding properties and esterase-like activity of the albumin mutants were nearly the same as those for the native albumin.4) The plasma levels of the albumin mutants labeled with ^<125>I after intravenous bolus injections to rats showed two-phase profiles but their half-lives were shorten than that for the native albumin.From the above results, there are potentials for the design of such albumin mutants but further investigations need to be carried out in order to serve the objectives of better safety and efficacy of albumin preparation.

本研究旨在利用定点诱变技术开发一种安全有效的白蛋白。结果表明：1)获得了K199A、K525A两个单突变体。重组蛋白折叠正确，在远紫外和近紫外CD光谱上显示出与天然白蛋白相似的模式，并且对针对天然白蛋白的多克隆抗血清具有反应性。2) GdnCl和DSC热变性实验表明，天然白蛋白和突变白蛋白的构象稳定性无显著差异。3)白蛋白突变体的配体结合特性和酯酶样活性与天然白蛋白几乎相同。4)大鼠静脉注射后，标记为^<125>I的白蛋白突变体血浆水平呈两相分布，但其半衰期比天然白蛋白短。从上述结果来看，设计这种白蛋白突变体是有潜力的，但为了更好地实现白蛋白制备的安全性和有效性，还需要进行进一步的研究。

项目成果

T.Sakai et al: "Interaction between indoxyl sulfate, a typical uremic toxin bound to site II, and ligands bound to site I of HSA"Pharmaceutical Research. 18(4)(in press). (2001)

T.Sakai 等人：“硫酸吲哚酚（一种与位点 II 结合的典型尿毒症毒素）与与 HSA 位点 I 结合的配体之间的相互作用”药物研究。

丸山徹,小田切優樹(分担)沢田康文 他(編集): "医薬品の適正使用のための臨床薬物動態"じほう. 834 (2000)

Toru Maruyama、Yuki Odagiri（撰稿人）、Yasufumi Sawada 等人（编辑）：“正确使用药物的临床药代动力学”Jiho 834 (2000)。

Y.Tsutsumi: "Interaction between two carboxylate endogeneous substances,bilirubin and an uremic toxin,CMPF,on human serum albumin" Pharmaceutical Research. 16(6)(in press). (1999)

Y.Tsutsumi：“两种羧酸内源性物质胆红素和尿毒症毒素 CMPF 对人血清白蛋白的相互作用”药物研究。

H.Watanabe et al: "Conformational stability and warfarin binding properties of human serum albumin studied by recombinant mutants"Biochemical Journal. (in press). (2001)

H.Watanabe 等人：“重组突变体研究的人血清白蛋白的构象稳定性和华法林结合特性”生物化学杂志。

Watanabe H, et al.: "Role of arg-410 and tyr-411 in human serum albumin for ligand binding and esterase-like activity."Biochem J.. 349(3). 813-9 (2000)

Watanabe H 等人：“arg-410 和 tyr-411 在人血清白蛋白中对于配体结合和酯酶样活性的作用。”Biochem J.. 349(3)。