Short Article Bivalent Tethering of SspB to ClpXP Is Required for Efficient Substrate Delivery: A Protein-Design Study

Summary studies indicate that substrate-delivery complexes con- tain just one SspB dimer and two ssrA-tagged sub- SspB homodimers deliver ssrA-tagged substrates to strates per ClpX hexamer (Wah et al., 2002), suggestingClpXP for degradation. SspB consists of a substrate that more than one dimer of SspB does not interact binding domain and an unstructured tail with a ClpX with ClpX.binding module (XB). Using computational design, we Does an SspB dimer with only a single XB module engineered an SspB heterodimer whose subunits did mediate efficient substrate delivery to ClpXP? To ad-not form homodimers. Experiments with the designed dress this question, we used computational design to molecule and variants lacking one or two tails demon- engineer SspB heterodimers with different numbers ofstrate that both XB modules are required for strong XB sequences and assayed these proteins for enhance- binding and efficient substrate delivery to ClpXP. As- ment of ClpXP degradation. SspB molecules with onlysembly of stable SspB-substrate-ClpX delivery com-