Nanomechanics of streptavidin hubs for molecular materials.
Nanomechanics of streptavidin hubs for molecular materials.
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DOI：
10.1002/adma.201103316
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发表时间：
2011-12-15
影响因子：
29.4
通讯作者：
Marszalek, Piotr E.
中科院分区：
文献类型：
--
作者：
Kim, Minkyu;Wang, Chien-Chung;Benedetti, Fabrizio;Rabbi, Mahir;Bennett, Vann;Marszalek, Piotr E.
关键词：
通过使用I27结构域来识别单分子测量，我们显然只选择了那些测量的张力超过I27，∼200pN的展开阈值力的原子力显微镜记录。因此，链霉亲和素四聚体在低于200 pN(如果有的话)的力量下破裂的任何事件都将被错过。为了确定在较低力下可能发生的破裂事件，我们使用第二个结构[(I27-SNase)3-SM]4进行力谱测量(图3a)。图3b显示了在[(I27-SNase)3-SM]4上获得的典型的AFM力延伸曲线。有趣的是，大约70%的单分子力延伸曲线包含4个(多达6个)SNase的特征展开力峰，记录在任何I27模块展开之前结束(图3b的上图)。大约30%的记录包含I27展开力峰(图3b的底板)，并且很少(大约5%的记录)它们显示出大于300pN的力。威力从未超过500Pn。和以前一样，我们分析了[(I27-SNase)3-SM]4的所有单分子记录中的最后一个展开力峰，并显示了它们的pdf(图3c中的黑色实线；直方图如图S8b所示)。我们还对没有SMS的SNase-I27结构进行了单分子力谱实验(图S7)，并在图3c中显示了分离力峰的pdf(灰色虚线)。令人惊讶的是，在100Pn左右，链霉亲和素pdf明显大于分离pdf。我们得出的结论是，这种差异的起源是由于发生在100pN附近的大量链霉亲和素破裂事件，这些事件在使用I27手柄的测量中被遗漏了。这些高和低破裂力的起源是什么？结构研究表明，相同的链霉亲和素单体(A、B、C和D)组装成四聚体作为二聚体[20，21](图4a)。根据链霉亲和素[20，21]的晶体结构，由于A和B(AB)和C和D(CD)界面上有较大接触面积(∼16 nm 2)的强相互作用，涉及17个氢键(H键)和其他非共价相互作用，单体与二聚体[21]紧密结合在一起。[45]单体A和D(AD)和B和C(BC)之间存在相对较弱的相互作用，只有两个H键和其他相互作用，涉及相对较小的接触面积(∼5 nm 2)。45]在接触面积小于∼2 nm 2的A和C以及B和D界面上发生了更弱的相互作用。[45]这种分析表明，链霉亲和素四聚体内的内部界面可能具有不同的机械强度，并且由于这种各向异性，链霉亲和素四聚体的机械强度实际上可能以一种复杂的方式依赖于拉力几何形状。综上所述，我们的AFM结果表明，当使用I27手柄时，我们主要探测AB或CD等链霉亲和素单体之间的破裂事件(图4)。这些事件可能断裂17个氢键，因此，需要在500 nm/S的拉动速度下超过300pN的巨大拉伸力。然而，当利用SNase的展开特征时，我们探索了链霉亲和素四聚体中发生在单体(AB或CD)和二聚体(AB和CD)之间的所有可能的断裂事件。后者可能只涉及4个氢键的断裂，因此，需要的拉伸力要低得多，约为100pN。我们只观察到几个高强度破裂事件的事实
By using I27 domains for the identification of single molecule measurements, we obviously selected only those AFM recordings in which the measured tension exceeded the unfolding threshold force of I27,∼ 200 pN. Therefore, any events in which streptavidin tetramers could break at forces lower than 200 pN (if any) would have been missed. To identify rupture events that could possibly occurred at lower forces, we used the second construct,[(I27-SNase) 3-SM] 4, for force spectroscopy measurements (Figure 3 a). Figure 3 b shows typical AFM force-extension curves obtained on [(I27-SNase) 3-SM] 4. Interestingly, around 70% of single molecule force-extension curves contained 4 (up to 6) characteristic unfolding force peaks of SNase with recordings ending before any I27 modules unfolded (top panel of Figure 3 b). About 30% of the recordings contained I27 unfolding force peaks (bottom panel of Figure 3 b) and very rarely (approximately 5% of recordings) they displayed a force greater than 300 pN. Never did the force exceed 500 pN. As before, we analyzed the last unfolding force peaks in all single molecule recordings of [(I27-SNase) 3-SM] 4 and showed their pdf (black solid line in Figure 3 c; the histograms are shown in Figure S8b). We also performed single molecule force spectroscopy experiments on the SNase-I27 construct without SMs (Figure S7) and show the pdf of detachment force peaks in Figure 3 c (gray dashed line). Surprisingly, at around 100 pN, the streptavidin pdf is significantly greater as compared to the detachment pdf. We conclude that the origin of this difference is caused by numerous streptavidin rupture events which occur at forces around 100 pN that were missed in measurements using I27 handles.What is the origin of these high and low rupture forces? Structural studies revealed that identical streptavidin monomers (A, B, C and D) are assembled into tetramers as dimers of dimers [20, 21](Figure 4a). Based on the crystal structure of streptavidin [20, 21], monomers are tightly associated into dimers [21] due to strong interactions across the A and B (AB) and C and D (CD) interfaces with large contact areas (∼ 16 nm 2), which involve 17 hydrogen bonds (H-bonds) and other noncovalent interactions.[45] Relatively weak interactions exist in cross-interfaces between monomers A and D (AD) and B and C (BC) with only two H-bonds and other interactions, involving relatively small contact areas (∼ 5 nm 2).[21, 45] Even weaker interactions occur across A and C and B and D interfaces with a contact area of less than∼ 2 nm 2.[45] This analysis suggests that the internal interfaces within a streptavidin tetramer may have different mechanical strengths and, because of this anisotropy, the mechanical strength of a streptavidin tetramer may actually depend, in a complicated fashion, on the pulling geometry. Taken together, our AFM results suggest that when using I27 handles, we mainly probed rupture events between streptavidin monomers such as AB or CD (Figure 4). These events likely rupture 17 H-bonds and therefore, require large stretching forces in excess of 300 pN at a pulling speed of 500 nm/s. However, when exploiting the unfolding signature of SNase, we probed all possible rupture events within the streptavidin tetramer that occur between monomers (AB or CD) and dimers (AB and CD). The latter likely involves the rupture of only 4 H-bonds and therefore, requires a significantly lower stretching force of approximately 100 pN. The fact that we only observed a few high force rupture events
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影响因子：
3.9
作者：
CHAIET, L;WOLF, FJ
通讯作者：
WOLF, FJ
影响因子：
56.9
作者：
Grubmuller, H;Heymann, B;Tavan, P
通讯作者：
Tavan, P
影响因子：
64.8
作者：
Lv, Shanshan;Dudek, Daniel M.;Li, Hongbin
通讯作者：
Li, Hongbin
影响因子：
56.9
作者：
LABEIT, S;KOLMERER, B
通讯作者：
KOLMERER, B
DOI：
10.1073/pnas.86.7.2190
发表时间：
1989-04-01
影响因子：
11.1
作者：
HENDRICKSON, WA;PAHLER, A;PHIZACKERLEY, RP
通讯作者：
PHIZACKERLEY, RP