您知道蛋白质折叠吗？这是一个很新的词。新到什么程度？您可以上网到著名的不列颠百科全书网站检索一下proteinfolding（即蛋白质折叠），还没有相应的解释。对于生命奥秘的探索，将贯穿新世纪乃至新千年人类的历史。而蛋白质如何形成三维空间结构，就是其中的一大课题。
很多生物过程通过形成有显著功能的中间体进行，蛋白质折叠也不例外。虽然鉴定和描述这种中间体可以为涉及的反应的机制提供关键的线索，但是由于中间体是瞬时存在的并且量也较少，获得这种中间体的信息便倍受挑战。
在7月29日出版的《自然》杂志上，多伦多大学生物化学系的Dmitry Korzhnev和他的导师Lewis Kay教授发表了一份对生物化学界来说意义重大的研究报告。在这份报告中，他们已经首次在原子水平上成功地绘制出“中间态蛋白质”三维结构的基础结构图。
他们借助核磁共振（nuclear magnetic resonance，NMR）技术鉴定了SH3的两个48位氨基酸突变体（G48M和G48V）的中间态，SH3来源于Fyn酪氨酸激酶，而此低量的中间态是通过未折叠和完全折叠两种状态的动态平衡得到的。他们在不同温度下，进行核磁共振，对折叠的过程以及中间态的结构进行了描述。
学术界认为该项研究成果为解开生物学界最重要的问题——关于蛋白质如何形成三维空间结构，给出了关键性的解答。
Low-populated folding intermediates of Fyn SH3 characterized by relaxation dispersion NMR
Many biochemical processes proceed through the formation of functionally significant intermediates. Although the identification and characterization of such species can provide vital clues about the mechanisms of the reactions involved, it is challenging to obtain information of this type in cases where the intermediates are transient or present only at low population. One important example of such a situation involves the folding behaviour of small proteins that represents a model for the acquisition of functional structure in biology. Here we use relaxation dispersion nuclear magnetic resonance (NMR) spectroscopy to identify, for two mutational variants of one such protein, the SH3 domain from Fyn tyrosine kinase, a low-population folding intermediate in equilibrium with its unfolded and fully folded states. By performing the NMR experiments at different temperatures, this approach has enabled characterization of the kinetics and energetics of the folding process as well as providing structures of the intermediates. A general strategy emerges for an experimental determination of the energy landscape of a protein by applying this methodology to a series of mutants whose intermediates have differing degrees of native-like structure.
Figure 3 Structural analysis of the I state of mutants G48M and G48V of Fyn SH3 using chemical shifts. a, Ratio of 15N chemical-shift differences, |
exp(k)| = |
FI(k)/FU(k)|, plotted on the ribbon trace of the structure of the wild-type Fyn SH3 domain27, with the colour code indicated. Residues with values of exp < -0.5 or exp > 2.5 are in grey. b, Comparison of exp (filled squares) with the calc values obtained in the structure determinations (lines); only residues with more than four native contacts have been represented. c, Ribbon representation of the ensemble of 25 structures determined for each mutant; the structure with the thicker ribbon is the more representative (lowest average pairwise r.m.s.d.) of the ensemble. d, Average effective energy contact map of the native state (bottom) and the ensembles obtained for the I state of each mutant (top); Eij is the interaction energy (in kcal mol-1) between the side chains of residues i and j according to the CHARMM19 force field. Residue numbers are indicated along each of the axes.
