Figure 5
Dimerization of protein D is essential for AMP binding. (a) Protein D is composed of protein A and B detected by non-reducing SDS-PAGE followed by Western blotting using antibodies raised against protein A (albumin, middle) and protein B (α1-antitrypsin, right). (b, c) Dimerization of protein D. The proteins A, B and D were cross-linked with glutaraldehyde (GA) and separated by 8% reduced SDS-PAGE (b). The cross-linked protein D was further analyzed by Western blotting using antibodies raised against protein A (albumin, middle) and protein B (α1-antitrypsin, right) (c). 1X GA represents 0.00625% glutaraldehyde. (d, e) Inhibition of dimerization of protein D by β-mercaptoethanol (β-ME) and SDS. The protein D was cross-linked with 0.025% glutaraldehyde in the presence of β-ME (d) or SDS (e). 1X represents 0.05% β-ME (d) and 0.0625% SDS (e), respectively. (f) SDS inhibit the pulled-down of protein D to immobilized TP4-A12I,A15I which analyzed by 15% non-reducing SDS-PAGE. 1X represents 0.0625% SDS.
