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. 2015 Sep 10;2(3):ofv116. doi: 10.1093/ofid/ofv116

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© The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America.

This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs licence (http://creativecommons.org/licenses/by-nc-nd/4.0/), which permits non-commercial reproduction and distribution of the work, in any medium, provided the original work is not altered or transformed in any way, and that the work is properly cited. For commercial re-use, please contact journals.permissions@oup.com.

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Figure 3. — Mapping of β-1,2-oligomannosides epitopes on yeast and hyphal mannoproteins. (A and B) Western blots of whole-cell extracts from different reference strains (CAI-4 and BWP17) and mutants (mnt1Δ/mnt2Δ and bmtsΔ) grown at 28°C were stained with anti-β-1,2-oligomannosides monoclonal antibody (mAb) 5B2 (A and B) and lectin concanavalin A (ConA), specific for terminal α-mannosides (A). Phosphopeptidomannan (PPM) and cell wall mannoproteins (CWMPs) are indicated. (C) Six His-tagged recombinant Hwp1 (6xHis-Hwp1) was produced and secreted in the culture supernatant by CAI-4 strain. Recombinant Hwp1 was purified on nickel column together with a zinc-binding protein, Pra1. The 2 purified hyphal mannoproteins were analyzed by Western blot with mAbs 16B1, specific for Hwp1, and 5B2. Before separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the proteins were treated or not with PNGase F. (D) The 6xHis-Hwp1 and Pra1 were expressed by CAI-4 strain and bmt1Δ mutant, purified on nickel column, and analyzed by Western blot with mAbs 16B1 and 5B2 and lectin ConA. The 2 proteins are indicated. All the mutants and their reference strains are Ura⁺ with CIp10 integrated in the same RPS1 allele.