Localization characteristics of cell wall synthesis protein Wag31 and penicillin binding protein C in Clavibacter michiganensis

CmWag31 is a member of the DivIVA family of proteins in <i>Clavibacter michiganensis</i>. The DivIVA family have been demonstrated to play a key role in the synthesis of cell wall peptidoglycan and cell division in most bacterial species. It has been previously confirmed that the <i>pbp</i>C (penicillin-binding protein C) deletion mutants affect bacterial division and cell wall synthesis. Based on the confirmation of the interaction between CmWag31 and CmPBPC, the present study conducted a systemic analysis on their localization characteristics. The results indicated that CmWag31 exhibited the capacity to interact with the transglycosylase (TG) and transpeptidase (TP) domain of CmPBPC, while CmPBPC only interacted with the NTD region of CmWag31. Co-localization analysis showed that CmWag31 co-localized with CmPBPC at the bacterial growth tips of <i>Clavibacter michiganensis</i> and <i>Escherichia coli</i>. The mutation of R19A, R19C, A99T, and A102T of CmWag31 resulted in abnormal localization in <i>Escherichia coli</i>. In the case of <i>C. michiganensis,</i> the CmWag31^A102T protein exhibited a diffuse localization, which is a departure from the polar localization of its wild type. The co-localization of the CmWag31^A102T mutation with CmPBPC exhibited discrepancies between <i>C. michiganensis</i> and <i>E. coli</i>. The diffused localization of CmWag31^A102T can be restored by overexpression of CmPBPC in <i>C. michiganensis</i>, yet this restoration is not observed in <i>E. coli</i>. This result indicates that CmPBPC from <i>C. michiganensis</i> may not fully excute their function in <i>E. coli</i> due to species-specific differences.