Asp2 and Asp3 bind the serine-rich repeat regions of GspB. (A) Schematic of GST-GspB736 and GST-GspB truncates containing specific GspB domains. (B) The GST-GspB fusion proteins were affinity purified and probed by far-Western blotting with purified H6Asp2 or H6Asp3. Lane 1, GST; lane 2, GST-GspB736; lane 3, GST-SP; lane 4, GST-AST; lane 5, GST-SRR1; lane 6, GST-BR; lane 7, GST-SRR2′. Each fusion protein was detected with anti-GST antibody (blot I). Binding of the Asps to the GST fusion proteins was detected with anti-H6 antibody (blots II and III). (C) Far-Western blotting assessing binding of the Asps to SRR1. Lane 1, purified Gsp736FLAG served as a positive control; lane 2, purified SP-AST-SRR1FLAG fusion protein; lane 3, purified SP-ASTFLAG fusion protein. The membranes were probed with purified H6Asp2 or H6Asp3, followed by detection of the bound Asp with anti-H6 antibody (blots II and III). The amount of GspB or fusion protein present in each lane was detected with anti-FLAG antibody (blot I).