Fuccio Carmelo, Luchinat Enrico, Barbieri Letizia, Neri Sara, Fragai Marco
Magnetic Resonance Center (CERM), University of Florence, Via L. Sacconi 6, 50019, Sesto Fiorentino, Florence, Italy.
Department of Biomedical, Clinical and Experimental Sciences, University of Florence, Viale Morgagni 50, 50134, Florence, Italy.
J Biomol NMR. 2016 Apr;64(4):275-80. doi: 10.1007/s10858-016-0026-0. Epub 2016 Apr 22.
In-cell NMR provides structural and functional information on proteins directly inside living cells. At present, the high costs of the labeled media for mammalian cells represent a limiting factor for the development of this methodology. Here we report a protocol to prepare a homemade growth medium from Spirulina platensis autolysate, suitable to express uniformly labeled proteins inside mammalian cells at a reduced cost-per-sample. The human proteins SOD1 and Mia40 were overexpressed in human cells grown in (15)N-enriched S. platensis algal-derived medium, and high quality in-cell NMR spectra were obtained.
细胞内核磁共振直接在活细胞内部提供有关蛋白质的结构和功能信息。目前，用于哺乳动物细胞的标记培养基的高成本是该方法发展的一个限制因素。在此，我们报告了一种从钝顶螺旋藻自溶物制备自制生长培养基的方案，该培养基适合在哺乳动物细胞内以降低的每个样品成本表达均匀标记的蛋白质。人类蛋白质超氧化物歧化酶1（SOD1）和Mia40在富含（15）N的钝顶螺旋藻藻类衍生培养基中生长的人类细胞中过表达，并获得了高质量的细胞内核磁共振光谱。
