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. 2019 Aug 8;9:11409. doi: 10.1038/s41598-019-47657-9

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© The Author(s) 2019

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Design and purification of the Ag85b (a), Sbi III-IV-Ag85b (b) and Sbi III-IV (c) constructs. Schematic representations of the constructs are shown on the left and SDS-PAGE profiles of the nickel affinity, ion exchange and size exclusion chromatography elution fractions are shown in the right panels. All three constructs bear an N-terminal 6x His-tag and the numbering of Ag85b and Sbi III-IV amino acid residues is according to their respective full-length proteins. The Sbi III-IV-Ag85b conjugate contains Sbi domains III and IV conjugated via a 28 amino acid histidine and glycine-rich linker to Ag85b at the C-terminus. Note: the yield of Ag85b shown is from 1 L of culture while the yield from Sbi III-IV-Ag85b and Sbi III-IV is from 4 L of culture. Details of the protein physiochemical properties can be found in Supplementary Table S1. Supplementary Fig. S1 includes purification chromatogram profiles. Full-length gel images of (a–c) are included in Supplementary Figs S11–S13.