2-DIMENSIONAL H-1-NMR STUDY OF RECOMBINANT INSECT DEFENSIN-A IN WATER - RESONANCE ASSIGNMENTS, SECONDARY STRUCTURE AND GLOBAL FOLDING
2-DIMENSIONAL H-1-NMR STUDY OF RECOMBINANT INSECT DEFENSIN-A IN WATER - RESONANCE ASSIGNMENTS, SECONDARY STRUCTURE AND GLOBAL FOLDING
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DOI：
10.1007/bf01875319
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发表时间：
1992-05-01
影响因子：
2.7
通讯作者：
ACHSTETTER, T
中科院分区：
文献类型：
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作者：
BONMATIN, JM;BONNAT, JL;ACHSTETTER, T
报道了重组昆虫防御素A的500 MHz 2D H-1核磁共振研究。这个40个残基的防御蛋白含有3个二硫键，带正电荷，具有抗菌特性。对重组防御素A的2D核磁共振图谱进行了全归属，并对其二级结构元素进行了定位。NH质子的NOE连接性、3J(NH-α-H)耦合常数以及H-1/H-2交换率和Delta-Delta/Delta-T温度系数有力地支持了α螺旋(残基14-24)和反平行β折叠(残基27-40)的存在。用DISMAN程序生成骨架折叠模型，用Amber程序精炼能量。这是在以下条件的基础上完成的。(I)133个选定的NOE，(Ii)21个来自3J(NH-α-H)偶合常数的二面体约束，(Iii)12个氢键，主要由H-1/H-2交换率或温度系数得出，此外还有9个初始二硫键共价约束。这两个二级结构元件和连接它们的两个弯曲涉及大约70%的残基总数，这使得分子的C-末端部分具有一定的稳定性。其余的N-末端片段形成一个定义不太清楚的环。这种空间结构，其中一个β-折叠通过两个二硫键连接到一个α-螺旋上，通过第三个二硫键连接到一个大环上，与蝎子轮藻毒素中发现的非常相似，似乎部分存在于几种无脊椎动物毒素中。
A 500 MHz 2D H-1 NMR study of recombinant insect defensin A is reported. This defense protein of 40 residues contains 3 disulfide bridges, is positively charged and exhibits antibacterial properties. 2D NMR maps of recombinant defensin A were fully assigned and secondary structure elements were localized. The set of NOE connectivities, 3J(NH-alpha-H) coupling constants as well as H-1/H-2 exchange rates and DELTA-delta/DELTA-T temperature coefficients of NH protons strongly support the existence of an alpha-helix (residues 14-24) and of an antiparallel beta-sheet (residues 27-40). Models of the backbone folding were generated by using the DISMAN program and energy refined by using the AMBER program. This was done on the basis of. (i) 133 selected NOEs, (ii) 21 dihedral restraints from 3J(NH-alpha-H) coupling constants, (iii) 12 hydrogen bonds mostly deduced from H-1/H-2 exchange rates or temperature coefficients, in addition to 9 initial disulfide bridge covalent constraints. The two secondary structure elements and the two bends connecting them involve approximately 70% of the total number of residues, which impose some stability in the C-terminal part of the molecule. The remaining N-terminal fragment forms a less well defined loop. This spatial organization, in which a beta-sheet is linked to an alpha-helix by two disulfide bridges and to a large loop by a third disulfide bridge, is rather similar to that found in scorpion charybdotoxin and seems to be partly present in several invertebrate toxins.