Sunday, May 21, 2006

Day 2 - Chemistry Session: Protein Biosynthesis: Encoding Unnatural Amino Acids and Evolving in Cells - Lei Wang

"Proteins are involved in virtually every life process." 20 amino acids are encoded by 61 of the 64 codons. To generate interesting protein properties, there are two avenues: to increase the building blocks (i.e. genetically encode unnatural amino acids), and 2. to permute and recombine the existing AAs (?)

1. Is it possible to expand the genetic code to include new amino acids? The Amino acids, aminoacyl-tRNA synthetase, tRNA, and codon all must have a high specificity to insure fidelity and minimize cross talk. They tried many different tRNA/aaRS pairs, finally settling on one already existing in M. jannaschii.

Here is a general approach for the generation and improvement of orthogonal tRNAs:
generated libraries of the synthetase to evolve tRNAs that both 1) work in

final product had translational fidelity near 99%, comprable to natural tRNAs.

Labeling in vitro?
Also could make homogenous glycoproteins (help understand glycan dymanics?)

(check nature article about redesigning firefly luciferase to be more red)

Replace group with heavy metal to facilitate x-ray structure determination?

Could incorporate photo-active groups for reporting certain activities, or metal chelators to manipulate electron transport chains, etc.

sequence variants of a 100-aa "protein" 20^100 = 10^130 > number of atoms in the universe (~10^80).

B-Cells encounter and become activated by antigen causes introduction of point mutations in V region of lg at a rate 100 times greater than baseline mutation rate.

protein evolution using SHM?

Ratio Sorting for Red-Shifted cells (using FACS?) Longer wavelengths have better tissue penetration.

This was fairly technical and I plan on tracking down Dr. Wang later for clarification.

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