Totally synthetic microperoxidase-11

Area of Science:

• Biochemistry
• Synthetic Chemistry
• Biomaterials

Background:

• Microperoxidase-11 (MP-11) is a heme-containing peptide with peroxidase activity.
• Natural MP-11 is typically obtained from cytochrome c, limiting its availability and modification.
• Developing synthetic routes for MP-11 is crucial for broader applications and mutant studies.

Purpose of the Study:

• To report the successful synthesis of a fully synthetic microperoxidase-11 (MP-11).
• To characterize the catalytic activity and electrochemical properties of the synthetic MP-11.
• To establish a convenient, nature-independent method for preparing MP-11 and its analogs.

Main Methods:

• Solid-phase peptide synthesis of the undecapeptide VQKCAQCHTVE.
• Thiol-ene click reaction for haemin reconstitution.
• High-speed atomic force microscopy (HS-AFM) for structural confirmation.
• Enzymatic assays (TMB oxidation) and electrochemical measurements (cyclic voltammetry).
• Computer simulation for electrocatalytic reaction parameter analysis.

Main Results:

• Successful synthesis and reconstitution of MP-11 using chemical methods.
• Synthetic MP-11 exhibited significant catalytic activity (27% of natural MP-11) in TMB oxidation.
• A point mutation (H8M) drastically reduced catalytic activity (2.7%).
• Electrochemical studies showed direct electron transfer and catalytic current-voltage behavior comparable to natural MP-11.
• Experimental data were well-reproduced by computer simulations, yielding consistent electrocatalytic parameters.

Conclusions:

• A totally synthetic MP-11 has been successfully prepared via solid-phase peptide synthesis and haemin reconstitution.
• The synthetic MP-11 displays comparable catalytic and electrochemical performance to the natural enzyme.
• This synthetic approach provides a versatile and reliable method for producing MP-11 and its mutants without relying on natural sources.