Solvent effects on the solubility and physical stability of human Insulin-like Growth Factor I
Solvent effects on the solubility and physical stability of human Insulin-like Growth Factor I
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DOI：
10.1023/a:1012101027814
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发表时间：
1997-05-01
影响因子：
3.7
通讯作者：
FlorinRobertsson, E
中科院分区：
文献类型：
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作者：
Fransson, J;Hallen, D;FlorinRobertsson, E
目的。研究了人胰岛素样生长因子I (hIGF-I)在不同赋形剂的水溶液中的溶解度和物理稳定性。方法。 hIGF-I 的溶解度通过紫外线吸收和遮光颗粒的定量来确定。采用差示扫描量热法 (DSC) 和圆二色性 (CD) 光谱研究了 hIGF-I 的物理稳定性。结果。人 IGF-I 在 140 mM 苯甲醇和 145 mM 氯化钠存在下低温沉淀。 CD 数据显示，与 5 mM 磷酸盐缓冲液中的相比，这些条件下 hIGF-I 的三级结构受到干扰。在苯甲醇存在下，290 mM 甘露醇稳定 hIGF-I。氯化钠或甘露醇本身对溶解度或三级结构没有影响。苯甲醇被 hIGF-I 吸引，而氯化钠被优先排除。氯化钠增强了苯甲醇的吸引力，导致 hIGF-I 盐析。 CD数据表明苯甲醇与hIGF-I中苯丙氨酸的相互作用。 hIGF-I 的热变性发生在所有含有氯化钠的溶液中，而甘露醇或苯甲醇对热稳定性没有影响。 hIGF-I 的热稳定性因此在 145 mM 氯化钠中降低，尽管它被排除在 hIGF-I 之外。结论。 hIGF-I 的自缔合和热聚集是由疏水相互作用驱动的。苯甲醇被 hIGF-I 吸引并诱导三级结构发生变化，从而导致蛋白质在低温下产生疏水性吸引力。
Purpose. The solubility and physical stability of human Insulin-like Growth Factor I (hIGF-I) were studied in aqueous solutions with different excipients.Methods. The solubility of hIGF-I was determined by UV-absorption and quantification of light blocking particles. The physical stability of hIGF-I was studied with differential scanning calorimetry (DSC) and circular dichroism (CD) spectroscopy.Results. Human IGF-I precipitated at low temperature in the presence of 140 mM benzyl alcohol and 145 mM sodium chloride. CD data showed that the tertiary structure of hIGF-I during these conditions was perturbed compared to that in 5 mM phosphate buffer. In the presence of benzyl alcohol 290 mM mannitol stabilized hIGF-I. Sodium chloride or mannitol by themselves had no effect on either the solubility or the tertiary structure. Benzyl alcohol was attracted to hIGF-I, whereas sodium chloride was preferentially excluded. The attraction of benzyl alcohol was reinforced by sodium chloride leading to salting-out of hIGF-I. The CD-data indicated interactions of benzyl alcohol with phenylalanine in hIGF-I. Thermal denaturation of hIGF-I occurred in all solutions with sodium chloride, whereas mannitol or benzyl alcohol had no effect on the thermal stability. The thermal stability of hIGF-I was thus decreased in 145 mM sodium chloride although it was excluded from hIGF-I.Conclusions. The self-association and thermal aggregation of hIGF-I is driven by hydrophobic interactions. Benzyl alcohol is attracted to hIGF-I and induces changes in the tertiary structure causing hydrophobic attraction of the protein at low temperatures.