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Publication Abstract from PubMed

Nanobodies or VHHs (Variable Heavy domains of Heavy chain) are single domain antibodies that comprise three antigenic complementary determining regions (CDR). Nanobodies are used in numerous scientific applications including, bio-imaging, diagnosis, therapeutics, and macromolecular crystallography. We obtained crystals of a approximately 14 kDa nanobody specific for the NEIL1 DNA glycosylase (hereafter called A5) in 0.5 M ammonium sulfate, 0.1 M sodium citrate tribasic dihydrate pH 5.6, and 1.0 M lithium sulfate monohydrate from the Crystal HT Hampton Research screen that were further optimized. Here, we describe the structure determination and refinement of the A5 crystals to a resolution of 2.1 A. The data collected were complicated by the presence of anisotropy and twinning, and while initial space group determination pointed to a higher apparent tetragonal crystal system, the data statistics suggested twinning, placing the crystal in an orthorhombic system. Twinning was confirmed by the Padilla and Yeates test, H-test, and Britton test based on local intensity differences with a twin fraction of 0.4. Molecular replacement produced the best solution in the orthorhombic space group P2 (1) 2 (1) 2 with four molecules in the asymmetric unit and we were able to model over 96% of the residues in the electron density with a final R (work) and R (free) of 0.1988 and 0.2289 upon refinement. SYNOPSIS: The crystal structure of a specific nanobody against NEIL1 was determined to 2.1 A. The structure was ultimately solved in an orthorhombic space group after diffraction data analysis revealed mild anisotropy as well as pseudo-merohedral twinning.

Deciphering the orthorhombic crystal structure of a novel NEIL1 nanobody with pseudo-merohedral twinning.,Thompson MK, Sharma N, Prakash A bioRxiv. 2023 Aug 7:2023.08.07.552313. doi: 10.1101/2023.08.07.552313. Preprint. PMID:37609231^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.