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Gonococcal penicillin-binding protein 3 and the surface-exposed 44kDa peptidoglycan-binding protein appear to be the
1Laboratories of Microbial Pathogenesis, Veterans Affairs Medical Center, Decatur, Georgia 30033.
Abstract:
The outer membrane of Neisseria gonorrhoeae contains a 44,000 dalton (44kDa) surface-exposed protein which has the reported ability to form covalent interactions with peptidoglycan (PG). This PG-binding outer-membrane protein (OMP) appears to be highly conserved since it has been detected in all isolates examined. It also appears to be invariant since its primary structure among strains gives evidence of being identical (Judd et al., 1991). While studying the interaction of gonococcal penicillin-binding proteins (PBPs) with human lysosomal cathepsin G, we noticed that the 44kDa PG-binding OMP exhibited certain properties similar to PBP3. In this study we sought to obtain biochemical evidence to ascertain whether these proteins were the same. We found that both proteins fractionated with other sarkosyl-insoluble OMPs and that they exhibited similar susceptibility to cleavage in situ by enzymatically active cathepsin G. Moreover, a purified preparation of the 44kDa OMP was found to covalently bind radiolabelled benzylpenicillin in vitro. Thus, the data presented herein suggest that the 44kDa PG-binding OMP and PBP3 are the same OMP.
Insights
Neisseria gonorrhoeae outer membrane protein (OMP) PBP3, a 44kDa protein, covalently binds to peptidoglycan (PG). This study provides biochemical evidence that the conserved 44kDa PG-binding OMP is identical to PBP3.
Area of Science:
- Microbiology
- Bacterial Outer Membrane Proteins
- Neisseria gonorrhoeae
Background:
- Neisseria gonorrhoeae possesses a conserved 44kDa outer membrane protein (OMP) that binds peptidoglycan (PG).
- This OMP exhibits invariant primary structure across strains.
- Initial observations suggested similarities between this 44kDa OMP and penicillin-binding protein 3 (PBP3).
Purpose of the Study:
- To determine if the 44kDa PG-binding OMP and PBP3 are the same protein.
- To provide biochemical evidence for the identity of these two proteins.
Main Methods:
- Fractionation of sarkosyl-insoluble outer membrane proteins.
- Assessment of susceptibility to cleavage by cathepsin G.
- In vitro binding assay with radiolabeled benzylpenicillin.
Main Results:
- Both the 44kDa OMP and PBP3 fractionated together in the sarkosyl-insoluble fraction.
- Both proteins demonstrated similar susceptibility to cathepsin G cleavage.
- Purified 44kDa OMP preparation showed in vitro covalent binding of benzylpenicillin.
Conclusions:
- The 44kDa PG-binding OMP and PBP3 are biochemically indistinguishable.
- The data strongly suggest that the conserved 44kDa PG-binding OMP is PBP3 in Neisseria gonorrhoeae.