Abstract
S-Adenosylmethionine (SAM) is a natural metabolite having important uses in the treatment of various diseases. To develop a simple and effective way to produce SAM, immobilized Escherichia coli cells highly expressing an engineered variant of methionine adenosyltransferase (MAT) were employed to synthesize SAM. The recombinant I303V MAT variant was successfully produced at approximately 900 mg/L in a 10-L bioreactor and exhibited significantly less product inhibition and had a four-fold higher specific activity (14.2 U/mg) than the wild-type MAT (3.6 U/mg). To reduce the mass transfer resistance, the free whole-cells were permeabilized and immobilized using gellan gum gel as support in the presence of 100 mg/L Fe3O4 nanoparticles, and the highest activity (4152.4 U/L support) was obtained, with 78.2% of the activity recovery. The immobilized cells were more stable than the free cells under non-reactive conditions, with a half-life of 9.1 h at 50 °C. Furthermore, the magnetically immobilized cells were employed to produce SAM at a 40-mM scale. The residual activity of the immobilized cells was 67% of its initial activity after 10 reuses, and the conversion rate of ATP was ≥95% in all 10 batches. These results indicated that magnetically immobilized cells should be a promising biocatalyst for the biosynthesis of SAM.
Highlights
The primary metabolite S-adenosylmethionine (SAM), which is synthesized in vivo from and adenosine triphosphate (ATP) by methionine adenosyltransferase (MAT), plays important roles in transsulfuration, transmethylation, and polyamine synthesis [1,2]
Among the assayed concentrations of magnetic Fe3 O4 nanoparticles, the enzyme activity for SAM synthesis was highest at a concentration of 100 mg/L, where 40 mM of ATP was activity for SAM synthesis was highest at a concentration of 100 mg/L, where 40 mM of ATP was completely converted to SAM in 4 h, while the equivalent amount of ATP could be completely converted completely converted to SAM in 4 h, while the equivalent amount of ATP could be completely in 5 h by nonmagnetically immobilized cells. These results revealed that SAM biosynthetic activity of converted in 5 h by nonmagnetically immobilized cells
The specific activity of I303V MAT had a four-fold increase compared to the recombinant wild-type MAT
Summary
The primary metabolite S-adenosylmethionine (SAM), which is synthesized in vivo from and adenosine triphosphate (ATP) by methionine adenosyltransferase (MAT), plays important roles in transsulfuration, transmethylation, and polyamine synthesis [1,2]. It is widely used in treating clinical disorders such as liver disease, osteoarthritis, and depressive syndromes [3,4,5]. Genetic modification of yeast cells and the optimization of fermentation conditions are the two primary approaches to improving the production of SAM [13,14,15,16,17,18].
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