期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2020
卷号：117
期号：11
页码：5844-5852
DOI：10.1073/pnas.1922264117
出版社：The National Academy of Sciences of the United States of America
摘要：Human profilin I reduces aggregation and concomitant toxicity of the polyglutamine-containing N-terminal region of the huntingtin protein encoded by exon 1 (htt ex1 ) and responsible for Huntington’s disease. Here, we investigate the interaction of profilin with htt ex1 using NMR techniques designed to quantitatively analyze the kinetics and equilibria of chemical exchange at atomic resolution, including relaxation dispersion, exchange-induced shifts, and lifetime line broadening. We first show that the presence of two polyproline tracts in htt ex1 , absent from a shorter huntingtin variant studied previously, modulates the kinetics of the transient branched oligomerization pathway that precedes nucleation, resulting in an increase in the populations of the on-pathway helical coiled-coil dimeric and tetrameric species (τex ≤ 50 to 70 μs), while leaving the population of the off-pathway (nonproductive) dimeric species largely unaffected (τex ∼750 μs). Next, we show that the affinity of a single molecule of profilin to the polyproline tracts is in the micromolar range ( K diss ∼ 17 and ∼ 31 μM), but binding of a second molecule of profilin is negatively cooperative, with the affinity reduced ∼11-fold. The lifetime of a 1:1 complex of htt ex1 with profilin, determined using a shorter huntingtin variant containing only a single polyproline tract, is shown to be on the submillisecond timescale ( τ ex ∼ 600 μs and K diss ∼ 50 μM). Finally, we demonstrate that, in stable profilin–htt ex1 complexes, the productive oligomerization pathway, leading to the formation of helical coiled-coil htt ex1 tetramers, is completely abolished, and only the pathway resulting in “nonproductive” dimers remains active, thereby providing a mechanistic basis for how profilin reduces aggregation and toxicity of htt ex1 .
