Logvinova D S, Nikolaeva O P, Levitsky D I
Bach Institute of Biochemistry, Research Center of Biotechnology, Russian Academy of Sciences, Moscow, 119071, Russia.
Biochemistry (Mosc). 2017 Feb;82(2):213-223. doi: 10.1134/S0006297917020134.
We applied dynamic light scattering (DLS) to compare aggregation properties of two isoforms of myosin subfragment 1 (S1) containing different "essential" (or "alkali") light chains, A1 or A2, which differ by the presence of an N-terminal extension in A1. Upon mild heating (up to 40°C), which was not accompanied by thermal denaturation of the protein, we observed a significant growth in the hydrodynamic radius of the particles for S1(A1), from ~18 to ~600-700 nm, whereas the radius of S1(A2) remained unchanged and equal to ~18 nm. Similar difference between S1(A1) and S1(A2) was observed in the presence of ADP. In contrast, no differences were observed by DLS between these two S1 isoforms in their complexes S1-ADP-BeF and S1-ADP-AlF which mimic the S1 ATPase intermediate states S1*-ATP and S1**-ADP-P. We propose that during the ATPase cycle the A1 N-terminal extension can interact with the motor domain of the same S1 molecule, and this can explain why S1(A1) and S1(A2) in S1-ADP-BeF and S1-ADP-AlF complexes do not differ in their aggregation properties. In the absence of nucleotides (or in the presence of ADP), the A1 N-terminal extension can interact with actin, thus forming an additional actin-binding site on the myosin head. However, in the absence of actin, this extension seems to be unable to undergo intramolecular interaction, but it probably can interact with the motor domain of another S1 molecule. These intermolecular interactions of the A1 N-terminus can explain unusual aggregation properties of S1(A1).
我们应用动态光散射（DLS）来比较肌球蛋白亚片段1（S1）的两种同工型的聚集特性，这两种同工型含有不同的“必需”（或“碱性”）轻链A1或A2，A1因在N端有一个延伸而与A2不同。在温和加热（高达40°C）时，蛋白质未发生热变性，我们观察到S1（A1）颗粒的流体动力学半径显著增大，从约18 nm增大到约600 - 700 nm，而S1（A2）的半径保持不变，约为18 nm。在存在ADP的情况下，S1（A1）和S1（A2）之间也观察到类似差异。相比之下，在模拟S1 ATP酶中间状态S1 * -ATP和S1 ** -ADP-P的复合物S1-ADP-BeF和S1-ADP-AlF中，通过DLS未观察到这两种S1同工型之间存在差异。我们提出，在ATP酶循环过程中，A1的N端延伸可以与同一S1分子的运动结构域相互作用，这可以解释为什么S1-ADP-BeF和S1-ADP-AlF复合物中的S1（A1）和S1（A2）在聚集特性上没有差异。在没有核苷酸的情况下（或存在ADP时），A1的N端延伸可以与肌动蛋白相互作用，从而在肌球蛋白头部形成一个额外的肌动蛋白结合位点。然而，在没有肌动蛋白的情况下，这个延伸似乎无法进行分子内相互作用，但它可能与另一个S1分子的运动结构域相互作用。A1 N端的这些分子间相互作用可以解释S1（A1）不寻常的聚集特性。
