The accessory helix of complexin functions by stabilizing central helix secondary structure.
The accessory helix of complexin functions by stabilizing central helix secondary structure.
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DOI：
10.7554/elife.04553
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发表时间：
2014-11-10
期刊：
影响因子：
7.7
通讯作者：
Dittman JS
中科院分区：
文献类型：
--
作者：
Radoff DT;Dong Y;Snead D;Bai J;Eliezer D;Dittman JS
突触前蛋白复合素(CPX)是突触小泡融合的重要调节因子，但其调控机制尚不清楚。其高度保守的中央螺旋(CH)直接与三元SNARE复合体结合，是所有已知CPX功能所必需的。相邻的辅助螺旋(AH)虽然在CPX功能中也起着重要作用，但并不保守，其机制已被提出了许多模型。我们利用线虫在体内和体外检测了AH突变和嵌合体对CPX功能的影响。当小鼠AH被替换为蠕虫CPX时，其功能完全恢复，表明其机制在进化上是保守的。当进入CH的螺旋传播中断时，CPX抑制功能受损，而用非天然螺旋序列取代AH可恢复CPX功能。我们认为，AH是通过稳定CH二级结构而不是通过蛋白质或脂类相互作用来发挥作用的。DOI：http://dx.doi.org/10.7554/eLife.04553.001，神经系统以电信号的形式在全身发送信息，这些信息通过被称为神经元的细胞传递。这些信号无法穿过隔开相邻神经元的小缝隙--称为突触。相反，当电信号到达突触时，称为神经递质的化学物质会穿过突触释放出来，并在下一个神经元中触发电信号。神经递质储存在神经元内的小囊膜中，称为突触小泡。当囊泡与神经元周围的膜融合时，它们就会释放出来。这种融合过程必须受到严格控制，以确保信息在正确的时间在神经元之间传递。复合蛋白是一种小蛋白，通过与一组称为SNARE复合体的蛋白质结合来控制囊泡融合。它包含两个有结构的部分，称为中央螺旋和副螺旋，这两个部分对囊泡融合都很重要。中央螺旋能够与SNARE蛋白结合，它在所有动物中都有相同的氨基酸序列-蛋白质的构建块。然而，副螺旋中的氨基酸序列在不同的动物中差异很大，目前还不清楚它在所有动物中是否都发挥着相同的作用。Radoff等人。对线虫体内的络合素进行了研究，发现用小鼠的氨基酸序列替换其副螺旋后，仍能正确地控制囊泡融合。事实上，当它的附属螺旋被形状相似的人造蛋白质螺旋取代时，它仍然可以正常工作。这些实验表明，辅助螺旋的整体结构比其确切的氨基酸序列更重要。Radoff等人。提出它在囊泡融合中的作用是稳定中央螺旋的结构，使其能够与SNARE蛋白结合。下一个挑战是了解当复杂蛋白与SNARE蛋白结合时如何阻止囊泡融合。DOI：http://dx.doi.org/10.7554/eLife.04553.002
The presynaptic protein complexin (CPX) is a critical regulator of synaptic vesicle fusion, but the mechanisms underlying its regulatory effects are not well understood. Its highly conserved central helix (CH) directly binds the ternary SNARE complex and is required for all known CPX functions. The adjacent accessory helix (AH) is not conserved despite also playing an important role in CPX function, and numerous models for its mechanism have been proposed. We examined the impact of AH mutations and chimeras on CPX function in vivo and in vitro using C. elegans. The mouse AH fully restored function when substituted into worm CPX suggesting its mechanism is evolutionarily conserved. CPX inhibitory function was impaired when helix propagation into the CH was disrupted whereas replacing the AH with a non-native helical sequence restored CPX function. We propose that the AH operates by stabilizing CH secondary structure rather than through protein or lipid interactions. DOI: http://dx.doi.org/10.7554/eLife.04553.001 The nervous system sends information around the body in the form of electrical signals that travel through cells called neurons. These signals cannot pass across the small gaps—called synapses—that separate neighboring neurons. Instead, when electrical signals reach the synapse, chemicals called neurotransmitters are released across the gap and trigger an electrical signal in the next neuron. Neurotransmitters are stored within neurons in small envelopes of membrane known as synaptic vesicles. They are released when the vesicles fuse with the membrane that surrounds the neuron. This fusion process must be tightly controlled to ensure that information is passed between the neurons at the right time. Complexin is a small protein that controls vesicle fusion by binding to a group of proteins called the SNARE complex. It contains two structured sections called the central helix and the accessory helix, which are both important for vesicle fusion. The central helix is able to bind to the SNARE proteins, and it has the same sequence of amino acids—the building blocks of proteins—in all animals. However, the sequence of amino acids in the accessory helix varies widely across different animals and it is not clear whether it performs the same role in all of them. Radoff et al. studied complexin in the nematode worm C. elegans, and found that when its accessory helix is replaced with the amino acid sequence from the mouse one, it can still properly control vesicle fusion. Indeed, complexin can still work properly when its accessory helix is replaced with an artificial protein helix that has a similar shape. These experiments suggest that the overall structure of the accessory helix is more important than its exact sequence of amino acids. Radoff et al. propose that its role in vesicle fusion is to stabilize the structure of the central helix to allow it to bind to the SNARE proteins. The next challenge is to understand how vesicle fusion is prevented when complexin binds to the SNARE proteins. DOI: http://dx.doi.org/10.7554/eLife.04553.002