کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
2803048 | 1156720 | 2009 | 6 صفحه PDF | دانلود رایگان |
![عکس صفحه اول مقاله: Insulin-like growth factor-I (IGF-I): Solution properties and NMR chemical shift assignments near physiological pH Insulin-like growth factor-I (IGF-I): Solution properties and NMR chemical shift assignments near physiological pH](/preview/png/2803048.png)
ObjectiveInsulin-like growth factor-I (IGF-I) plays important roles in normal growth and development, as well as in disease states, and its structure and function have been studied extensively using nuclear magnetic resonance (NMR) spectroscopy. However, IGF-I typically gives poor quality NMR spectra containing many broad peaks, because of aggregation at the protein concentrations generally required for NMR experiments as well as the internal dynamics of the molecule. The present study was undertaken to determine a reliable set of assignments under more physiological conditions.DesignSeveral reports of chemical shift assignments have been published previously for IGF-I either bound to a ligand or at relatively low pH (∼3–4), but there are many contradictions among them, reflecting the poor behaviour of IGF-I. Low pH conditions are also suboptimal for the analysis of interactions between IGF-I and IGF binding proteins (IGFBP) or IGFBP fragments. Spectra were recorded at low concentrations in order to identify conditions of temperature and pH where all peaks could be observed.ResultsWe show that good quality 2D 1H–15N HSQC spectra of 15N-labelled IGF-I can be obtained at pH 6 and 37 °C, much closer to physiological conditions, by using lower IGF-I concentrations (0.05 mM). Surprisingly, at this concentration and temperature, spectra were of better quality at pH 6 than at pH 4, in contrast to previous observations made at millimolar concentrations of IGF-I. We were then also able to assign the chemical shifts of IGF-I at pH 6 and 37 °C using 3D heteronuclear spectra recorded on a 0.7 mM 15N/13C-labelled IGF-I sample.ConclusionThese results provide a valuable resource for future studies of the structure, dynamics, folding, and binding interactions of IGF-I, as well as analogues thereof, by means of NMR spectroscopy.
Journal: Growth Hormone & IGF Research - Volume 19, Issue 3, June 2009, Pages 226–231