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LytU-SH3b fusion protein as a novel and efficient enzybiotic against methicillin-resistant Staphylococcus aureus
Mortaza Taheri-Anganeh1, Seyyed Hossein Khatami2, Zeinab Jamali3
1Department of Medical Biotechnology, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran.
Abstract:
Methicillin-resistant Staphylococcus aureus (MRSA) is a challenging infectious agent worldwide. The ever growing antibiotic resistance has made the researchers to look for new anti-staphylococcal agents. Autolysins are staphylococcal enzymes that lyse bacterial cell wall for cell division. Autolysins can be used as novel enzybiotics (enzymes have antibiotic effects) for staphylococcal infections. LytU is a newly explored autolysin. SH3b is a potent cell wall binding domain that can be fused to lytic enzymes to increase their activity. The aim of this study was to design a novel and efficient fusion enzybiotic that could lyse staphylococcal cell wall peptidoglycan by disrupting the bacteria. LytU-SH3b fusion construct was synthesized and LytU was amplified through the construct, using overhang PCR. The fusion and native forms that had his-tag were synthesized by recombinant technology in Escherichia coli BL21 (DE3) strain and purified utilizing Ni-NTA agarose beads. LytU and LytU-SH3b activity and potency were assessed using plate lysis assay, turbidity reduction assay and minimal inhibitory concentration (MIC) tests. All these tests showed that LytU-SH3b has more activity and potency than LytU. LytU-SH3b has MIC 421 fold lesser than LytU. Finally, LytU-SH3b is a novel and efficient recombinant enzybiotic that can lyse MRSA as an alternative to chemical small molecule antibiotics.
Insights
Researchers developed a novel fusion enzybiotic, LytU-SH3b, to combat Methicillin-resistant Staphylococcus aureus (MRSA). This engineered enzyme demonstrates enhanced potency in lysing MRSA cell walls, offering a promising alternative to traditional antibiotics.
Area of Science:
- Microbiology
- Biotechnology
- Enzymology
Background:
- Methicillin-resistant Staphylococcus aureus (MRSA) poses a significant global health challenge due to increasing antibiotic resistance.
- Novel therapeutic strategies, including enzybiotics, are crucial for treating staphylococcal infections.
- Autolysins, bacterial cell wall-degrading enzymes, and cell wall binding domains like SH3b are key components for developing effective enzybiotics.
Purpose of the Study:
- To design and characterize a novel fusion enzybiotic, LytU-SH3b, for enhanced lysis of staphylococcal cell walls.
- To evaluate the activity and potency of the LytU-SH3b fusion protein compared to the native LytU enzyme.
- To explore the potential of LytU-SH3b as an alternative therapeutic agent against MRSA infections.
Main Methods:
- LytU-SH3b fusion construct synthesized using overhang PCR.
- Recombinant expression of LytU and LytU-SH3b in Escherichia coli BL21 (DE3) with subsequent purification using Ni-NTA agarose beads.
- Assessment of enzyme activity and potency via plate lysis assay, turbidity reduction assay, and minimal inhibitory concentration (MIC) tests.
Main Results:
- LytU-SH3b exhibited significantly higher lytic activity and potency compared to the native LytU enzyme.
- The minimal inhibitory concentration (MIC) of LytU-SH3b was 421-fold lower than that of LytU, indicating superior efficacy.
- Successful synthesis and purification of the recombinant LytU-SH3b fusion protein.
Conclusions:
- LytU-SH3b is a novel and highly effective recombinant enzybiotic with potent MRSA cell wall lytic capabilities.
- The fusion of the SH3b domain to LytU enhances its activity, making it a promising alternative to conventional antibiotics.
- This study highlights the potential of engineered enzybiotics as a new class of antimicrobial agents against resistant bacteria.
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