AI Article Synopsis
- Researchers investigated how varying levels of fluorinated amino acids affect the function and stability of an enzyme called chloramphenicol acetyltransferase (CAT).
- They found that replacing all leucine residues with 5',5',5'-trifluoroleucine (TFL) retains enzymatic activity at room temperature and improves secondary structure, but reduces stability when exposed to heat and solvents.
- While the enzyme's catalytic activity remains similar to the wild type, increasing TFL levels leads to decreased stability, indicating that enhanced structure from fluorination doesn’t guarantee better overall stability.
Article Abstract
Varied levels of fluorinated amino acid have been introduced biosynthetically to test the functional limits of global substitution on enzymatic activity and stability. Replacement of all the leucine (LEU) residues in the enzyme chloramphenicol acetyltransferase (CAT) with the analog, 5',5',5'-trifluoroleucine (TFL), results in the maintenance of enzymatic activity under ambient temperatures as well as an enhancement in secondary structure but loss in stability against heat and denaturants or organic co-solvents. Although catalytic activity of the fully substituted CAT is preserved under standard reaction conditions compared to the wild-type enzyme both in vitro and in vivo, as the incorporation levels increase, a concomitant reduction in thermostability and chemostability is observed. Circular dichroism (CD) studies reveal that although fluorination greatly improves the secondary structure of CAT, a large structural destabilization upon increased levels of TFL incorporation occurs at elevated temperatures. These data suggest that enhanced secondary structure afforded by TFL incorporation does not necessarily lead to an improvement in stability.
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| http://dx.doi.org/10.1002/bit.20910 | DOI Listing |
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