Isolation, biochemical characterization and anti-bacterial activity of BPIFA2 protein
Vladimir Prokopovic1, Milica Popovic1, Uros Andjelkovic2
1Faculty of Chemistry, Department of Biochemistry, University of Belgrade, Studentski trg 12-16, 11000 Belgrade, Serbia.
Objective:
Human BPIFA2 (parotid secretory protein) is a ubiquitous soluble salivary protein, which belongs to the PLUNC family of proteins. Having sequence similarity to bactericidal/permeability-increasing protein and lipopolysaccharide-binding protein, PLUNC proteins are probably involved in local antibacterial response at mucosal sites, such as oral cavity. The aim of the study was to isolate and characterize human BPIFA2.
Design:
In this paper, we report one-step affinity chromatography method for BPIFA2 purification from whole human saliva. The isolated BPIFA2 was identified by trypsin mass fingerprinting and characterized by electrophoretic methods. Antibacterial activity of BPIFA2 against model microorganism Pseudomonas aeruginosa was shown in minimum inhibitory concentration and time kill study assays.
Results:
The protein showed microheterogeneity, both in molecular weight and pI value. BPIFA2 inhibited the growth of P. aeruginosa in microgram concentration range determined by minimum inhibitory concentration assay. In the time kill study, 32μg/mL BPIFA2 showed clear bactericidal activity and did not cause any aggregation of bacteria.
Conclusion:
Affinity chromatography is well suited for isolation of functional BPIFA2 with a potent bactericidal activity against P. aeruginosa.
Insights
Human BPIFA2 (parotid secretory protein) exhibits potent antibacterial activity against Pseudomonas aeruginosa. This study successfully isolated and characterized BPIFA2 from saliva, demonstrating its effectiveness in combating bacterial growth.
Area of Science:
- Biochemistry
- Microbiology
- Salivary Proteins
Background:
- Human BPIFA2, also known as parotid secretory protein, is a salivary protein in the PLUNC family.
- PLUNC proteins share similarities with bactericidal/permeability-increasing protein and lipopolysaccharide-binding protein, suggesting a role in mucosal antibacterial defense.
- The oral cavity, a mucosal site, is constantly exposed to microorganisms, necessitating robust local defense mechanisms.
Purpose of the Study:
- To isolate and characterize human BPIFA2 (parotid secretory protein).
- To investigate the antibacterial activity of purified BPIFA2 against Pseudomonas aeruginosa.
Main Methods:
- One-step affinity chromatography was employed for BPIFA2 purification from human saliva.
- Protein identification was confirmed using trypsin mass fingerprinting.
- Electrophoretic methods were used for characterization, and antibacterial activity was assessed via minimum inhibitory concentration and time-kill assays.
Main Results:
- Purified BPIFA2 displayed microheterogeneity in molecular weight and pI.
- BPIFA2 demonstrated significant inhibition of P. aeruginosa growth in the microgram concentration range.
- A concentration of 32μg/mL BPIFA2 exhibited clear bactericidal activity without causing bacterial aggregation.
Conclusions:
- One-step affinity chromatography is an effective method for isolating functional BPIFA2.
- Human BPIFA2 possesses potent bactericidal activity against Pseudomonas aeruginosa.
- BPIFA2 represents a promising component of the oral innate immune system.
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