C. K. Sahu, V. K. Bayineni, J. S. Hampapura, C. M. Hussain, R.-K. Kadeppagari
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Characterization of 36-kDa Dimeric Methotrexate-degrading Enzyme from Variovorax paradoxus, its Performance and Stability of its Cross-linked Aggregates
In the present study, methotrexate (MTX)-degrading 36-kDa dimer was characterized from the 0–80% ammonium sulphate precipitate of Variovorax paradoxus cell lysate. This enzyme showed optimum activity at 35°C and pH 6.5 with KM value of 199.6 µM for MTX which are different from those of 46-kDa dimer. Micro- or nano-sized cross-linked aggregates of new enzyme showed better stability in the presence of serum than the native soluble form. Addition of an endogenic carrier protein, human serum albumin (HSA) to the enzyme aggregates further improved their serum stability. Nanoaggregates showed better serum stability over microaggregates. Nanoaggregates of the enzyme degraded the MTX faster than soluble form and microaggregates due to their lower KM values on MTX, whereas microaggregates were slower than soluble enzyme itself due to their higher KM values. Hence, cross-linked aggregates of MTX- degrading enzyme isolated from V. paradoxus shown better performance in their nanoform compared to the microform. Nanoaggregates of enzyme revealed the highest functionality and serum stability that makes them more suitable for therapeutic applications.
期刊介绍：
Applied Biochemistry and Microbiology is an international peer reviewed journal that publishes original articles on biochemistry and microbiology that have or may have practical applications. The studies include: enzymes and mechanisms of enzymatic reactions, biosynthesis of low and high molecular physiologically active compounds; the studies of their structure and properties; biogenesis and pathways of their regulation; metabolism of producers of biologically active compounds, biocatalysis in organic synthesis, applied genetics of microorganisms, applied enzymology; protein and metabolic engineering, biochemical bases of phytoimmunity, applied aspects of biochemical and immunochemical analysis; biodegradation of xenobiotics; biosensors; biomedical research (without clinical studies). Along with experimental works, the journal publishes descriptions of novel research techniques and reviews on selected topics.
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