Synthesis and Incorporation of Unnatural Amino Acids To Probe and Optimize Protein Bioconjugations.
The utilization of unnatural amino acids (UAAs) in bioconjugations is ideal due to their ability to confer a degree of bioorthogonality and specificity. In order to elucidate optimal conditions for the preparation of bioconjugates with UAAs, we synthesized 9 UAAs with variable methylene tethers (2-4) and either an azide, alkyne, or halide functional group. All 9 UAAs were then incorporated into green fluorescent protein (GFP) using a promiscuous aminoacyl-tRNA synthetase. The different bioconjugations were then analyzed for optimal tether length via reaction with either a fluorophore or a derivatized resin. Interestingly, the optimal tether length was found to be dependent on the type of reaction. Overall, these findings provide a better understanding of various parameters that can be optimized for the efficient preparation of bioconjugates.
Department of Chemistry, College of William & Mary P.O. Box 8795, Williamsburg, Virginia 23187, United States.
Johnathan C
JC
Maza
Department of Chemistry, College of William & Mary P.O. Box 8795, Williamsburg, Virginia 23187, United States.
Jaclyn R
JR
McKenna
Department of Chemistry, College of William & Mary P.O. Box 8795, Williamsburg, Virginia 23187, United States.
Benjamin K
BK
Raliski
Department of Chemistry, College of William & Mary P.O. Box 8795, Williamsburg, Virginia 23187, United States.
Matthew T
MT
Freedman
Department of Chemistry, College of William & Mary P.O. Box 8795, Williamsburg, Virginia 23187, United States.
Douglas D
DD
Young
D000480:Alkynes / Q000737:chemistry; D000596:Amino Acids / Q000138:chemical synthesis* / Q000737:chemistry*; D001386:Azides / Q000737:chemistry; D060326:Chemistry Techniques, Synthetic; D049452:Green Fluorescent Proteins / Q000737:chemistry*; D006219:Halogens / Q000737:chemistry; D008958:Models, Molecular; D017433:Protein Structure, Secondary
D000480:Alkynes; D000596:Amino Acids; D001386:Azides; D006219:Halogens; D049452:Green Fluorescent Proteins