Yang Shi-Zhen, Huang Yi-Ding, Jie Xin-Feng, Feng You-Min, Niu Jing-Yi
World J Gastroenterol. 2000 Jun;6(3):371-373. doi: 10.3748/wjg.v6.i3.371.
To study the relationship between insulin A chain regions and insulin biological activities, we designed a series of insulin analogues with changes at A21, A12-18 of C terminal helical region and A8-10 located in the region of A6-A11 intra-chain disulphide bond.METHODS:Insulin A-chain analogues were prepared by stepwise Fmoc solid phase manual synthesis and then combined with natural B-chain of porcine insulin to yield corresponding insulin analogues. Their biological activities were tested by receptor binding, mouse convulsion and immunological assay.RESULTS: A21Ala Ins retains 70.3% receptor binding capacity and 60% in vivo biological activity.DesA13-14, A21Ala Ins and DesA12-13-14-15, A21Ala Ins still have definite biological activity,7.9% and 4.0% receptor binding,and 6.2% and 3.3% in vivo biological activity respectively. A15Asn, A17Pro, A21Ala Ins maintains 10.4% receptor binding and 10% in vivo biological activity. A8His, A9Arg, A10Pro, A21Ala Ins, A8His, A9Lys, A10Pro, A21Ala Ins and A8His, A9Lys, A10Arg, A21Ala Ins have 51.9%, 44.3% and 32.1% receptor binding respectively,50%, 40% and 30% in vivo biological activity respectively, and 28.8%, 29.6% and 15.4% immunological activity respectively.CONCLUSION:A21Asn can be replaced by simple amino acid residues.The A chains with gradually damaged structural integrity in A12-18 helical region and the demolition of the A12-18 helical region by the substitution of Pro and Asn for A17Glu and A15Gln respectively can combine with the B chain and the combination products show definite biological activity, the helical structure of A12-18 is essential for biological activities of insulin. A8-10 is not much concerned with biological activities, but is much more important antigenically in binding to its antibodies, these results may help us design a new type of insulin analogue molecule.
为研究胰岛素A链区域与胰岛素生物学活性之间的关系，我们设计了一系列胰岛素类似物，其在A21、C末端螺旋区域的A12 - 18以及位于A6 - A11链内二硫键区域的A8 - 10处有变化。方法：通过逐步Fmoc固相手动合成制备胰岛素A链类似物，然后与猪胰岛素的天然B链结合以产生相应的胰岛素类似物。通过受体结合、小鼠惊厥和免疫测定来测试它们的生物学活性。结果：A21Ala Ins保留70.3%的受体结合能力和60%的体内生物学活性。缺失A13 - 14、A21Ala Ins和缺失A12 - 13 - 14 - 15、A21Ala Ins仍具有一定的生物学活性，受体结合率分别为7.9%和4.0%，体内生物学活性分别为6.2%和3.3%。A15Asn、A17Pro、A21Ala Ins保持10.4%的受体结合率和10%的体内生物学活性。A8His、A9Arg、A10Pro、A21Ala Ins、A8His、A9Lys、A10Pro、A21Ala Ins和A8His、A9Lys、A10Arg、A21Ala Ins的受体结合率分别为51.9%、44.3%和32.1%，体内生物学活性分别为50%、40%和30%，免疫活性分别为28.8%、29.6%和15.4%。结论：A21Asn可被简单氨基酸残基取代。A12 - 18螺旋区域结构完整性逐渐受损的A链以及分别用Pro和Asn取代A17Glu和A15Gln导致A12 - 18螺旋区域被破坏的A链可与B链结合，且结合产物显示出一定的生物学活性，A12 - 18的螺旋结构对胰岛素的生物学活性至关重要。A8 - 10与生物学活性关系不大，但在与抗体结合方面抗原性更强，这些结果可能有助于我们设计新型胰岛素类似物分子。
