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. Author manuscript; available in PMC: 2009 Oct 5.
Published in final edited form as: Nat Cell Biol. 2008 Feb 10;10(3):272–282. doi: 10.1038/ncb1689

Figure 6. Dependence of N-glycan recognition for XTP3-B/OS-9 interaction with ERAD components and substrate.

Figure 6

a. Transient expression of S-tagged wild-type, mutant (R207A, R428A, R207A/R428A, N195Q, G379S) and a C-terminal truncation (Δ404-483) of XTP3-B in HEK293 cells, processed as in Fig. 4d and probed for Hrd1, SEL1L and S-tag by western blot. b. Expression of S-tagged wild type and MRH mutant (R188A) of OS-9.1. Immunoblots were performed with antibodies against S-tag, SEL1L and GRP94. c. NHK-HA coexpression with MRH mutants of XTP3-B (R207A/R428A) and OS-9.1 (R188A). Complexes were brought down with anti-HA and immunoblots probed with anti-S-tag (top) and anti-HA (bottom). Hairline indicates where western blot exposures were joined. Full scans are presented in Supplemental Information (Fig. S5). d. HA-tagged transthyretin (TTR-HA) wild-type (WT), D18G and A25T were coexpressed in HEK293 cells with S-tagged XTP3-B and OS-9.1. Complexes were affinity purified by S-protein agarose (top) or anti-HA (bottom) from cells lysed in 1% TritonX-100. Immunoblots were performed with anti-S-tag and anti-HA (TTR). Non-specific background bands are indicated by (*).