Pneumococcal CbpD is a murein hydrolase that requires a dual cell envelope binding specificity to kill target cells during fratricide.
Pneumococci that are competent for natural genetic transformation express a number of proteins involved in binding, uptake, translocation and recombination of DNA. In addition, they attack and lyse non-competent sister cells present in the same environment. This phenomenon has been termed fratricide. The key effector of pneumococcal fratricide is CbpD, a secreted protein encompassing an N-terminal CHAP domain, two SH3b domains and a C-terminal choline-binding domain (CBD). CbpD is believed to degrade the cell wall of target cells, but experimental evidence supporting this hypothesis has been lacking. Here, we show that CbpD indeed has muralytic activity, and that this activity requires functional CBD and SH3b domains. To better understand the critical role played by the non-catalytic C-terminal region of CbpD, various translational fusions were constructed between the CBD and SH3b domains and green fluorescent protein (GFP). The results showed that the SH3b domains specifically recognize and bind peptidoglycan, while the CBD domain functions as a localization signal that directs CbpD to the septal region of the pneumococcal cell. Intriguingly, transmission electron microscopy analysis revealed that target cells attacked by CbpD ruptures at the septal region, in accordance with the binding specificity displayed by the CBD domain.
Eldholm
Vegard
V
Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N-1432, As, Norway.
Johnsborg
Ola
O
Straume
Daniel
D
Ohnstad
Hilde Solheim
HS
Berg
Kari Helene
KH
Hermoso
Juan A
JA
Håvarstein
Leiv Sigve
LS
D000595:Amino Acid Sequence; D002473:Cell Wall / Q000378:metabolism*; D002794:Choline / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000235:genetics / Q000378:metabolism*; D010457:Peptidoglycan / Q000378:metabolism; D013296:Streptococcus pneumoniae / Q000201:enzymology* / Q000235:genetics / Q000648:ultrastructure; D018909:src Homology Domains
D010457:Peptidoglycan; D009238:N-Acetylmuramoyl-L-alanine Amidase; D002794:Choline