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Proteins, Expression, Isolation and Analysis> | The lysates were centrifuged at 12,000 × g for 15 min to remove cellular debris and the supernatants were incu- bated for 2 h with either Ni-NTA or glutathione resins (GenScript) at 4 °C with gentle rotation. | Get A Quote |
Posttranslational modification of key host proteins by virulence factors is an important theme in bacterial pathogenesis. A remarkable example is the reversible modifications of the small GTPase Rab1 by multiple effectors of the bacterial pathogen . Previous studies have shown that the effector SetA, dependent on a functional glucosyltransferase domain, interferes with host secretory pathways. However, the enzymatic substrate(s) of SetA in host cells remains unknown. Here, by using cross-linking mass spectrometry we uncovered Rab1 as the target of SetA during infection. Biochemical studies establish that SetA covalently attaches a glucose moiety to Thr within the switch II region of Rab1, inhibiting ... More