Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2024 Jul 1;25(8):3348–3372. doi: 10.1038/s44319-024-00183-w

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2024

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. Creative Commons Public Domain Dedication waiver http://creativecommons.org/publicdomain/zero/1.0/ applies to the data associated with this article, unless otherwise stated in a credit line to the data, but does not extend to the graphical or creative elements of illustrations, charts, or figures. This waiver removes legal barriers to the re-use and mining of research data. According to standard scholarly practice, it is recommended to provide appropriate citation and attribution whenever technically possible.

PMC Copyright notice

Figure EV4 — (A) Cartoon representation of the crystal structure of Mis18α_C-term/Mis18β_C-term (PDB ID: 7SFY). Mis18α is shown in purple and Mis18β in light pink. Potential residues involved in the interaction are highlighted. Mis18α (purple) and Mis18β (light pink). Right panel shows SDS-PAGE analysis of cobalt and amylose pull-down of His-MBP-Mis18β_{188–229 WT} with His-SUMO-Mis18α_191–233 mutants. SDS-PAGE shows protein bound to nickel resin as input (I) and protein-bound to amylose resin to assess interaction (P). Control with WT proteins shown in Fig. 4A. (B) Western blot analysis of co-immunoprecipitation (Co-IP) experiments using Mis18α antibody to test interaction of mCherry as a control, Mis18α−mCherry with and without mutations in the C-terminal α-helices and Mis18β-GFP. Top panel shows blot against mCherry, middle panel shows blot against GFP, and bottom panel shows blot against tubulin as loading control. (C) SEC-MALS of His-SUMO-Mis18α_{188-233 WT}, His-SUMO-Mis18α_{188-233 I201A/L205A} and His-SUMO-Mis18α_{188-233 L212A/L215A/L219A}. Normalised absorption at 280 nm (mAU, left y-axis) and molecular mass (kDa, right y-axis) are plotted against elution volume (ml, x-axis). Measured molecular weight (MW) and the calculated subunit stoichiometry based on the predicted MW. Samples were analysed using a Superdex 75 increase in 50 mM HEPES pH 8.0, 150 mM NaCl and 1 mM TCEP. (D) Representative immunoblots showing expression levels of endogenous proteins after treatment with siRNA. (E) Representative immunoblots showing expression levels of transiently expressed tagged proteins after transfection.