Krähenbühl Barbara, Wider Gerhard
ETH Zurich, Institute of Molecular Biology & Biophysics, Schafmattstrasse 30, CH-8093 Zürich, Switzerland.
Chimia (Aarau). 2012;66(10):770-4. doi: 10.2533/chimia.2012.770.
High resolution nuclear magnetic resonance (NMR) spectroscopy in solution is an established technique in structural biology. Detailed functional and structural studies of biological macromolecules by NMR require the assignment of the chemical shifts to specific nuclei. In biological applications, the necessary data is usually obtained from a number of two- and three-dimensional (2D and 3D) NMR experiments. Often, these data cannot be fully analyzed by automated computer programs due to insufficient separation and resolution of the signals in the available spectra. Then, complete resonance assignment requires manual interaction and can become a long and labor-intensive task. Automated projection spectroscopy (APSY) allows the substantial improvement of the resolution by providing spectral information from four and higher dimensional experiments without measuring the full spectrum, which would by far exceed any acceptable measuring time. APSY only measures a series of projections of the high-dimensional spectrum which can be obtained in a much shorter time. Peak picking of the projection spectra provides the basis for the calculation of the high-dimensional chemical shift correlation space by the algorithm GAPRO. The resulting high-dimensional peak lists are commonly artifact-free and of exceptional precision. Along with their high number of correlated nuclei they provide an ideal basis for reliable automated assignment. We will introduce the basic concepts of APSY, illustrate them with an application of a 6D APSY-seq-HNCOCANH experiment, and discuss some practical aspects.
溶液中的高分辨率核磁共振（NMR）光谱学是结构生物学中的一项成熟技术。通过NMR对生物大分子进行详细的功能和结构研究需要将化学位移分配到特定原子核上。在生物学应用中，必要的数据通常从多个二维和三维（2D和3D）NMR实验中获得。通常，由于可用光谱中信号的分离和分辨率不足，这些数据无法通过自动化计算机程序进行全面分析。因此，完整的共振归属需要人工干预，并且可能成为一项漫长且劳动强度大的任务。自动投影光谱学（APSY）通过提供来自四维及更高维实验的光谱信息而无需测量完整光谱，从而显著提高分辨率，因为测量完整光谱将远远超过任何可接受的测量时间。APSY仅测量高维光谱的一系列投影，这些投影可以在更短的时间内获得。投影光谱的峰挑选为通过GAPRO算法计算高维化学位移相关空间提供了基础。所得的高维峰列表通常无伪峰且精度极高。连同它们大量的相关原子核，它们为可靠的自动归属提供了理想的基础。我们将介绍APSY的基本概念，通过一个6D APSY-seq-HNCOCANH实验的应用来说明它们，并讨论一些实际问题。
