Effects of Non-Natural Amino Acid Incorporation into the Enzyme Core Region on Enzyme Structure and Function

International Journal of Molecular Sciences

H. Edward Wong¹  Inchan Kwon¹ 

[1] Department of Chemical Engineering, University of Virginia, Charlottesville, VA 22904, USA; E-Mail:

关键词: non-natural amino acids;    enzyme engineering;    enzyme kinetics;    site-specific incorporation;   

DOI  :  10.3390/ijms160922735

来源: mdpi

【 摘 要 】

Techniques to incorporate non-natural amino acids (NNAAs) have enabled biosynthesis of proteins containing new building blocks with unique structures, chemistry, and reactivity that are not found in natural amino acids. It is crucial to understand how incorporation of NNAAs affects protein function because NNAA incorporation may perturb critical function of a target protein. This study investigates how the site-specific incorporation of NNAAs affects catalytic properties of an enzyme. A NNAA with a hydrophobic and bulky sidechain, 3-(2-naphthyl)-alanine (2Nal), was site-specifically incorporated at six different positions in the hydrophobic core of a model enzyme, murine dihydrofolate reductase (mDHFR). The mDHFR variants with a greater change in van der Waals volume upon 2Nal incorporation exhibited a greater reduction in the catalytic efficiency. Similarly, the steric incompatibility calculated using RosettaDesign, a protein stability calculation program, correlated with the changes in the catalytic efficiency.

【 授权许可】

CC BY   
© 2015 by the authors; licensee MDPI, Basel, Switzerland.