Macromolecular assembly of bioluminescent protein nanoparticles for enhanced imaging.
Macromolecular assembly of bioluminescent protein nanoparticles for enhanced imaging.
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DOI：
10.1016/j.mtbio.2022.100455
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发表时间：
2022-12-15
影响因子：
8.2
通讯作者：
Wang, Szu-Wen
中科院分区：
文献类型：
--
作者：
Li, Enya;Brennan, Caroline K.;Ramirez, Aaron;Tucker, Jo A.;Butkovich, Nina;Meli, Vijaykumar S.;Ionkina, Anastasia A.;Nelson, Edward L.;Prescher, Jennifer A.;Wang, Szu-Wen
生物发光成像具有优于荧光成像的优点，例如最小的光漂白和自发荧光，以及在许多复杂环境中更大的信噪比。虽然在荧光素酶工程中已经取得了显著的成就，以产生明亮和稳定的报告，荧光素酶的纳米粒子跟踪的全部能力还没有得到全面的检查。在生物催化中，已经报道了固定在纳米颗粒上后增强的酶性能。因此，我们假设通过将荧光素酶组装到纳米颗粒上，所得复合物可以显着改善成像性能。使用模块化生物缀合策略，我们将NanoLuc（NLuc）或Akaluc生物发光蛋白连接到蛋白质纳米颗粒平台（E2），产生纳米颗粒NLuc-E2和Akaluc-E2，两者的直径均为45 nm。虽然在涉及Akaluc和Akaluc-E2的不同条件之间没有观察到显著差异，但pH 5.0下的游离NLuc显示出比pH 7.4下的游离NLuc显著更低的发射值。有趣的是，固定在E2纳米颗粒上的NLuc（NLuc-E2）在pH 7.4下发射增加的发光，并且在pH 5.0下显示出超过两个数量级（>200倍）的发光（比游离NLuc），扩大了使用纳米颗粒进行成像检测的潜力，甚至在内吞摄取时。在被巨噬细胞摄取后，在48 h时，使用NLuc-E2纳米颗粒产生的发光比使用游离NLuc高7倍。用NLuc-E2孵育的细胞也可以使用活体生物发光显微镜成像。最后，使用NLuc-E2通过成像检测纳米颗粒在淋巴结中的生物分布，但不使用常规标记的荧光E2。我们的数据表明，NLuc-bound纳米颗粒具有有利的特性，可用于从单细胞成像到体内生物分布的应用。
Bioluminescence imaging has advantages over fluorescence imaging, such as minimal photobleaching and autofluorescence, and greater signal-to-noise ratios in many complex environments. Although significant achievements have been made in luciferase engineering for generating bright and stable reporters, the full capability of luciferases for nanoparticle tracking has not been comprehensively examined. In biocatalysis, enhanced enzyme performance after immobilization on nanoparticles has been reported. Thus, we hypothesized that by assembling luciferases onto a nanoparticle, the resulting complex could lead to substantially improved imaging properties. Using a modular bioconjugation strategy, we attached NanoLuc (NLuc) or Akaluc bioluminescent proteins to a protein nanoparticle platform (E2), yielding nanoparticles NLuc-E2 and Akaluc-E2, both with diameters of ∼45 nm. Although no significant differences were observed between different conditions involving Akaluc and Akaluc-E2, free NLuc at pH 5.0 showed significantly lower emission values than free NLuc at pH 7.4. Interestingly, NLuc immobilization on E2 nanoparticles (NLuc-E2) emitted increased luminescence at pH 7.4, and at pH 5.0 showed over two orders of magnitude (>200-fold) higher luminescence (than free NLuc), expanding the potential for imaging detection using the nanoparticle even upon endocytic uptake. After uptake by macrophages, the resulting luminescence with NLuc-E2 nanoparticles was up to 7-fold higher than with free NLuc at 48 h. Cells incubated with NLuc-E2 could also be imaged using live bioluminescence microscopy. Finally, biodistribution of nanoparticles into lymph nodes was detected through imaging using NLuc-E2, but not with conventionally-labeled fluorescent E2. Our data demonstrate that NLuc-bound nanoparticles have advantageous properties that can be utilized in applications ranging from single-cell imaging to in vivo biodistribution.
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影响因子：
7.3
作者：
Brune KD;Howarth M
通讯作者：
Howarth M
影响因子：
46.2
作者：
Behzadi S;Serpooshan V;Tao W;Hamaly MA;Alkawareek MY;Dreaden EC;Brown D;Alkilany AM;Farokhzad OC;Mahmoudi M
通讯作者：
Mahmoudi M
影响因子：
5.6
作者：
Endo M;Ozawa T
通讯作者：
Ozawa T
影响因子：
12.6
作者：
Hu YB;Dammer EB;Ren RJ;Wang G
通讯作者：
Wang G
影响因子：
2.2
作者：
Harrell, Maria I.;Iritani, Brian M.;Ruddell, Alanna
通讯作者：
Ruddell, Alanna