Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, 113-8656, Tokyo, Japan.
Department of Applied Chemistry, Graduate School of Engineering, Kyushu University, 744 Moto-oka, 819-0395, Fukuoka, Japan.
Chembiochem. 2022 Nov 18;23(22):e202200476. doi: 10.1002/cbic.202200476. Epub 2022 Oct 26.
Methods for intracellular protein photoactivation have been studied to elucidate the spatial and temporal roles of proteins of interest. In this study, an intracellular protein photoactivation method was developed using sterically bulky caging. The protein of interest was modified with biotin via a photocleavable linker, and then conjugated with streptavidin to sterically block the protein surface for inactivation. The caged protein was transduced into cells and reactivated by light-induced degradation of the conjugates. A cytotoxic protein, saporin, was caged and photoactivated both in vitro and in living cells with this method. This method achieved control of the cytotoxic activity in an off-on manner, introducing cell death selectively at the designed location using light. This simple and versatile photoactivation method is a promising tool for studying spatio-temporal cellular events that are related to intracellular proteins of interest.
已经研究了细胞内蛋白质光激活的方法，以阐明感兴趣的蛋白质的空间和时间作用。在这项研究中，开发了一种使用大位阻笼蔽的细胞内蛋白质光激活方法。通过光可裂解的连接子将感兴趣的蛋白质修饰为生物素，然后与链霉亲和素缀合，以空间位阻方式使蛋白质表面失活。将笼蔽蛋白转导到细胞中，并通过光诱导缀合物的降解来重新激活。使用该方法，将细胞毒素蛋白蓖麻毒素进行笼蔽和光激活，无论是在体外还是在活细胞中都能实现。这种方法以开-关方式控制细胞毒性活性，使用光选择性地在设计的位置引入细胞死亡。这种简单而多功能的光激活方法是研究与感兴趣的细胞内蛋白质相关的时空细胞事件的有前途的工具。
