Pilotto, S. et al. Interplay among nucleosomal DNA, histone tails, and corepressor CoREST underlies LSD1-mediated H3 demethylation. Proc. Natl. Acad. Sci. 112, 2752–2757 (2015). It's built in slices which are connected to the one below by four studs around the central pillar. This enables them to be attached at angles other than 90 and 45 degrees, which helps give the finished conifer an organic appearance. Abeliovich, A. & Hammond, R. Midbrain dopamine neuron differentiation: Factors and fates. Dev. Biol. 304, 447–454 (2007). Kristensen, L. H. et al. Studies of H3K4me3 demethylation by KDM5B/Jarid1B/PLU1 reveals strong substrate recognition in vitro and identifies 2,4-pyridine-dicarboxylic acid as an in vitro and in cell inhibitor. FEBS J. 279, 1905–1914 (2012). We have previously raised nanobodies against KDM5B 22. One of these (NB8) was shown to bind strongly in SEC experiments whereas another (NB17) did not bind at all. To verify the structural integrity of the immobilized KDM5B, the positive control NB8 and the negative control NB17 were injected in a one-step gradient over immobilized KDM5B (881 RU) (Fig. 3E,F). NB8 showed a very strong binding to the KDM5B, with a slow off-rate. The binding was unbreakable using 1 M NaCl as regeneration solution and any suitable condition to break the interaction for regeneration of the immobilized surface was not found, wherefore experiments with multiple cycles were not possible. In comparison, NB17 did not show any specific binding to KDM5B. The gradient one-step injection of NB8 was fitted to a 1-1 interaction model deriving an approximate affinity of binding to KDM5B in higher pM range (Table S3). HDX-MS analysis of KDM5B

Svergun, D. I. Determination of the regularization parameter in indirect-transform methods using perceptual criteria. J. Appl. Crystallogr. 25, 495–503 (1992).

What's in the box

Nie, Y., Bergendahl, V., Hei, D. J., Jones, J. M. & Palecek, S. P. Scalable culture and cryopreservation of human embryonic stem cells on microcarriers. Biotechnol. Prog. 25, 20–31 (2009). Based on the observed FOXA2 expression patterns, we hypothesized that a desirable ventral fate was established and maintained more effectively in the 3D platform. To investigate this possibility, we examined the expression of additional ventral markers, SHH and CORIN, and found they were established in both platforms by D25, had dropped significantly by D40 in 2D, but were robustly maintained in 3D ( Figure S5). In summary, the gene expression patterns obtained here – including FOXA2 and LMX1A (floorplate derived midbrain fate), (ii) TFF3 (substantia nigra specific transcription factor), (iii) PITX3, GIRK2, NURR1, and TH (mature DA markers) – indicate that the cells differentiated in 3D acquired a substantia nigra specific mDA neuronal phenotype faster and in many cases to a greater extent than on 2D, while closely resembling the anticipated trends of mDA development schematically depicted in Fig. 1e 25, 26, 27, 28. Protein Structure & Function Programme, Macromolecular Crystallography Group, Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Blegdamsvej 3B, Copenhagen, 2200, Denmark

The price for the kit is a bit on the expensive side, but the ability to wire as one sees fit is a slight bonus. Kimanius, D., Forsberg, B. O., Scheres, S. & Lindahl, E. Accelerated cryo-EM structure determination with parallelisation using GPUs in RELION-2. bioRxiv 059717, https://doi.org/10.1101/059717 (2016). Jensen, P. F. et al. Investigating the Interaction between the Neonatal Fc Receptor and Monoclonal Antibody Variants by Hydrogen/Deuterium Exchange Mass Spectrometry. Mol. Cell. Proteomics 14, 148–161 (2015). Chen, V. C. et al. Scalable GMP compliant suspension culture system for human ES cells. Stem Cell Res. 8, 388–402 (2012). During modeling of a large number of truncations of KDM5B that comprise the PLU region, the alignment algorithm of the RaptorX server consistently identified actin molecules as templates for homology modelling. The HDX-MS data is a valuable validation source of the constructed models in this context. Remarkably, Fig. 6B with the mapping of the HDX-MS data on the model of the region show good agreement between the spectrin helical sections and slow HDX. A schematic representation of a hypothetical domain structure of KDM5B, including the FLD, is shown in Fig. 1. The FLD could not be modeled by RaptorX so interactions and the relative directionalities of the helices are undetermined from the HDX experiments only; consequently the helices are just drawn in a sequential manner.Biostructural Research, Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, Jagtvej 162, 2100, Copenhagen, Denmark Hughes, C. S., Postovit, L. M. & Lajoie, G. A. Matrigel: a complex protein mixture required for optimal growth of cell culture. Proteomics 10, 1886–1890 (2010). McNeish, J. et al. High-throughput screening in embryonic stem cell-derived neurons identifies potentiators of alpha-amino-3-hydroxyl-5-methyl-4-isoxazolepropionate-type glutamate receptors. J. Biol. Chem. 285, 17209–17 (2010).

Berrow, N. S. et al. A versatile ligation-independent cloning method suitable for high-throughput expression screening applications. Nucleic Acids Res. 35, e45 (2007). Khetan, S. et al. Degradation-mediated cellular traction directs stem cell fate in covalently crosslinked three-dimensional hydrogels. Nat. Mater. 12, 458–65 (2013). Veenvliet, J. V. J. et al. Specification of dopaminergic subsets involves interplay of En1 and Pitx3. Development 140, 3373–84 (2013). All stem cell procedures and procedures in animals were performed following NIH guidelines for animal care and use and were approved by the UC Berkeley Animal Care and Use Committee (ACUC), the Committee for Laboratory and Environmental Biosafety (CLEB), and the Stem Cell Research Oversight committee (SCRO).Oh, S. et al. Combined Nurr 1 and Foxa 2 roles in the therapy of Parkinson’s disease. EMBO Mol. Med. 7, 1–17 (2015).