Genetic encoding of a highly photostable, long lifetime fluorescent amino acid for imaging in mammalian cells.

Acridonylalanine (Acd) is a fluorescent amino acid that is highly photostable, with a high quantum yield and long fluorescence lifetime in water. These properties make it superior to existing genetically encodable fluorescent amino acids for monitoring protein interactions and conformational changes through fluorescence polarization or lifetime experiments, including fluorescence lifetime imaging microscopy (FLIM). Here, we report the genetic incorporation of Acd using engineered pyrrolysine tRNA synthetase (RS) mutants that allow for efficient Acd incorporation in both and mammalian cells.

Access to Faster Eukaryotic Cell Labeling with Encoded Tetrazine Amino Acids.

Labeling of biomolecules in live eukaryotic cells has been limited by low component stability and slow reaction rates. We show that genetically encoded tetrazine amino acids in proteins reach reaction rates of 8 × 10 M s with sTCO reagents, making them the fastest site-specific bioorthogonal labels in eukaryotic systems. We demonstrate that tetrazine amino acids are stable on proteins and are capable of quantitative labeling with sTCO reagents.

Growth Hormone Deficiency Causing Micropenis: Lessons Learned From a Well-Adjusted Adult.

This report of a 46,XY patient born with a micropenis consistent with etiology from isolated congenital growth hormone deficiency is used to (1) raise the question regarding what degree testicular testosterone exposure to the central nervous system during fetal life and early infancy has on the development of male gender identity, regardless of gender of rearing; (2) suggest the obligatory nature of timely full disclosure of medical history; (3) emphasize that virtually all 46,XY infants with functional testes and a micropenis should be initially boys except some with partial androgen insensitivity syndrome; and (4) highlight the sustaining value of a positive long-term relationship with a trusted physician (R.M.B.

Site-Specific Protein Labeling with Tetrazine Amino Acids.

Genetic code expansion is commonly used to introduce bioorthogonal reactive functional groups onto proteins for labeling. In recent years, the inverse electron demand Diels-Alder reaction between tetrazines and strained trans-cyclooctenes has increased in popularity as a bioorthogonal ligation for protein labeling due to its fast reaction rate and high in vivo stability. We provide methods for the facile synthesis of a tetrazine containing amino acid, Tet-v2.