Structure of CbpA J-Domain Bound to the Regulatory Protein CbpM Explains Its Specificity and Suggests Evolutionary Link between CbpM and Transcriptional Regulators

Abstract

CbpA is one of the six E. coli DnaJ/Hsp40 homologues of DnaK co-chaperones and the only one that is additionally regulated by a small protein CbpM, conserved in γ-proteobacteria. CbpM inhibits the co-chaperone and DNA binding activities of CbpA. This regulatory function of CbpM is accomplished through reversible interaction with the N-terminal J-domain of CbpA, which is essential for the interaction with DnaK. CbpM is highly specific for CbpA and does not bind DnaJ despite the high degree of structural and functional similarity between the J-domains of CbpA and DnaJ. Here we report the crystal structure of the complex of CbpM with the J-domain of CbpA. CbpM forms dimers and the J-domain of CbpA interacts with both CbpM subunits. The CbpM-binding surface of CbpA is highly overlapping with the CbpA interface for DnaK, providing a competitive model for regulation through forming mutually exclusive complexes. The structure also provides the explanation for the strict specificity of CbpM for CbpA, which we confirmed by making mutants of DnaJ that became regulated by CbpM. Interestingly, the structure of CbpM reveals a striking similarity to members of the MerR family of transcriptional regulators, suggesting an evolutionary connection between the functionally distinct bacterial co-chaperone regulator CbpM and the transcription regulator HspR.

DNA recognition by Escherichia coli CbpA protein requires a conserved arginine–minor-groove interaction

“…Structural information is not available for the intact CbpA protein. However, data are available for the J-domain, a CTDI–CTDII fragment, and the J-domain in complex with CbpM ( 9 – 11 ). Hence, CbpM forms a dimer that can bind two copies of the CbpA J-domain (Figure 1B i).…”

“…In combination with the structural studies described above molecular genetics has been used to reveal precise CbpA interaction surfaces ( 7 , 12 ). A basic patch on the surface of the J-domain, comprising amino acids R26, R30 and H33, interacts with CbpM side chains E64, T75 and E62 ( 11 , 12 ). Strikingly, this is the same surface of CbpA that contacts DnaK ( 11 ).…”

The Helicobacter pylori HspR-Modulator CbpA Is a Multifunctional Heat-Shock Protein

“…In other words, our data suggest that HspR can be regarded as a CbpA modulator. In E. coli, the gene lying immediately downstream cbpA codes for a protein modulator of CbpA activities called CbpM [12,13]. Interestingly, the crystal structure of CbpM reveals a strong similarity to the members of the MerR family of transcription regulators, to which HspR belongs [27].…”

“…Interestingly, the crystal structure of CbpM reveals a strong similarity to the members of the MerR family of transcription regulators, to which HspR belongs [27]. The region of the highest similarity between CbpM and MerR-like regulators is represented by the N-terminal portion, where is confined the DNA-binding domain of the transcriptional regulators [13]. In conclusion, data presented in this work support the idea of a reciprocal functional modulation between CbpA and HspR, an interaction that links HspR-mediated heat-shock gene regulation with CbpA-dependent protein folding and nucleoid maintenance, and stimulates further studies to better characterize this interplay.…”

“…This protein regulator is encoded by the gene laying immediately downstream cbpA within the same operon. The CbpA regulator CbpM essentially works as an inhibitor, forming a dimer able to bind two copies of the CbpA J-domain [12,13].…”