7sai

Crystal Structure of Lag30 Nanobody bound to eGFP

spinqualitylabelsall models 7sai, resolution 2.23Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 7sai is a 2 chain structure with sequence from Aequorea victoria and Lama glama. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 2.23Å Ligands: CRO, GOL, MES, PO4 Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function GFP_AEQVI Energy-transfer acceptor. Its role is to transduce the blue chemiluminescence of the protein aequorin into green fluorescent light by energy transfer. Fluoresces in vivo upon receiving energy from the Ca(2+)-activated photoprotein aequorin. Publication Abstract from PubMed Mammalian cell lines are important expression systems for large proteins and protein complexes, particularly when the acquisition of post-translational modifications in the protein's native environment is desired. However, low or variable transfection efficiencies are challenges that must be overcome to use such an expression system. Expression of recombinant proteins as a fluorescent protein fusion enables real-time monitoring of protein expression, and also provides an affinity handle for one-step protein purification using a suitable affinity reagent. Here, we describe a panel of anti-GFP and anti-mCherry nanobody affinity matrices and their efficacy for purification of GFP/YFP or mCherry fusion proteins. We define the molecular basis by which they bind their target proteins using X-ray crystallography. From these analyses, we define an optimal pair of nanobodies for purification of recombinant protein tagged with GFP/YFP or mCherry, and demonstrate these nanobody-sepharose supports are stable to many rounds of cleaning and extended incubation in denaturing conditions. Finally, we demonstrate the utility of the mCherry-tag system by using it to purify recombinant human topoisomerase 2alpha expressed in HEK293F cells. The mCherry-tag and GFP/YFP-tag expression systems can be utilized for recombinant protein expression individually or in tandem for mammalian protein expression systems where real-time monitoring of protein expression levels and a high-efficiency purification step is needed. High-efficiency recombinant protein purification using mCherry and YFP nanobody affinity matrices.,Cong ATQ, Witter TL, Schellenberg MJ Protein Sci. 2022 Sep;31(9):e4383. doi: 10.1002/pro.4383. PMID:36040252 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See Also Antibody 3D structures Green Fluorescent Protein 3D structures

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