During the previous funding cycle, we determined that a UAG-codon, located in the M. barkeri monomethylamine methyltransferase, encoded a new amino acid. Based on its electron density in the native and hydroxylamine complexes, we determined that the amino acid is 4-methyl-(4R,5R)-pyrroline-5- carboxylate, attached to the epsilon nitrogen of lysine. Based on its composition, we named this amino acid, L-pyrrolysine. The subsequent identification, by the Krzycki group, of an amber decoding tRNA(Pyl), and a novel L-pyrrolysyl-tRNA synthetase PylS, that loads L-pyrrolysine onto tRNA(Pyl) make a strong case for Lpyrrolysine being the 22nd genetically-encoded amino acid in nature. During this funding cycle, we propose to continue our efforts in understanding L-pyrrolysine's functional role and its mechanism of incorporation into proteins. This will include: (1) structural studies of L-pyrrolysine containing proteins, (2) structural studies involved in L-pyrrolysine incorporation into protein, and (3) chemical synthesis of L-pyrrolysine and its analogs.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM061796-08
Application #
7270620
Study Section
Macromolecular Structure and Function B Study Section (MSFB)
Program Officer
Fabian, Miles
Project Start
2000-08-05
Project End
2009-07-31
Budget Start
2007-08-01
Budget End
2008-07-31
Support Year
8
Fiscal Year
2007
Total Cost
$276,420
Indirect Cost
Name
Ohio State University
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
832127323
City
Columbus
State
OH
Country
United States
Zip Code
43210
Lee, Marianne M; Fekner, Tomasz; Lu, Jia et al. (2014) Pyrrolysine-inspired protein cyclization. Chembiochem 15:1769-72
Lee, Marianne M; Fekner, Tomasz; Tang, Tsz-Ho et al. (2013) A click-and-release pyrrolysine analogue. Chembiochem 14:805-8
Fekner, Tomasz; Chan, Michael K (2011) Hidden in plain sight: the biosynthetic source of pyrrolysine revealed. Chembiochem 12:2117-9
Fekner, Tomasz; Chan, Michael K (2011) The pyrrolysine translational machinery as a genetic-code expansion tool. Curr Opin Chem Biol 15:387-91
Li, Xin; Fekner, Tomasz; Chan, Michael K (2010) N6-(2-(R)-propargylglycyl)lysine as a clickable pyrrolysine mimic. Chem Asian J 5:1765-9
Fekner, Tomasz; Li, Xin; Lee, Marianne M et al. (2009) A pyrrolysine analogue for protein click chemistry. Angew Chem Int Ed Engl 48:1633-5
Li, Wen-Tai; Mahapatra, Anirban; Longstaff, David G et al. (2009) Specificity of pyrrolysyl-tRNA synthetase for pyrrolysine and pyrrolysine analogs. J Mol Biol 385:1156-64
Li, Xin; Fekner, Tomasz; Ottesen, Jennifer J et al. (2009) A pyrrolysine analogue for site-specific protein ubiquitination. Angew Chem Int Ed Engl 48:9184-7
Isaza, Clara E; Zhong, Xuejun; Rosas, Lucia E et al. (2008) A proposed role for Leishmania major carboxypeptidase in peptide catabolism. Biochem Biophys Res Commun 373:25-9
Lee, Marianne M; Bundschuh, Ralf; Chan, Michael K (2008) Distant homology detection using a LEngth and STructure-based sequence Alignment Tool (LESTAT). Proteins 71:1409-19

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