Charles Carter, Jr., PhD

RESEARCH INTERESTS: 

 
Protein Crystallography, Structural Polymorphism and Function

Life is driven by a harmonious interplay of structural polymorphism and ligand binding. To understand this interplay, we first seek accurate structural data concerning conformational variations. We have solved ground-state, activated pre-transition state, and product complex structures along the reaction pathways of tryptophanyl-tRNA synthetase, TrpRS, and cytidine deaminase, CDA. With atomic coordinates in hand, we then probe underlying structural features using biochemical and computational methods, working out mechanisms that link structural changes with recognition and catalysis. Many questions can best be addressed by combining structural and computational approaches. Collaborations with Jan Hermans and Alex Tropsha use molecular dynamics simulations, water-binding and likelihood potentials; others with Weitao Yang at Duke use quantum mechanics to map pathways connecting different structures along the TrpRS and CDA structural reaction profiles.

TrpRS, and its cognate tRNA^Trptogether translate the genetic code for tryptophan. Ground-state ligand-free TrpRS and its complexes with tryptophan and ATP are very open. Binding of both substrates induces a significant conformational change in which the ribose moiety of ATP is clamped into place for the activation reaction. In ternary complexes formed with ATP and non reactive tryptophan analogs, the compact enzyme conformation differs from both the "open" and "product-state" conformations. ATP binding drives this "induced fit". The quality of nonpolar sidechain packing is nonuniform within TrpRS; the highest-quality hydrophobic core apparently communicates the status of the active-site chemistry to a remote site that recognizes the tRNA^Trpanticodon. Prokaryotic TrpRSs differ considerably from eukaryotic counterparts, but are very similar to prokaryotic TyrRSs, making both enzymes good targets for structure-based, anti-infective drug discovery. We are currently using low angle X-ray scattering, steady-state kinetics, and isothermal titration calorimetry to study how amino acid and ATP binding affect the enzyme conformation, and its affinity for inhibitors.

Different binding geometries of CDA ligands representing substrate, transition state and product show that the enzyme "pulls" the 4NH₂ group and the pyrimidine ring of substrate cytidine in opposite directions. Electrostatic charge on the hydrolytic water molecule is apparently "buffered" by changes in a zinc-Sg bond distance, facilitating proton transfer from the substrate water to the leaving ammonia. Symmetry-breaking quaternary conformational changes are probably involved in opening and closing the CDA active site before and after catalysis, and are apparently are coupled to the binding and release of water molecules between domains. Sequence homologies between CDA and the cytidine deaminase subunit involved in apolipoprotein B messenger RNA editing suggest similar features in the editing complex, which apparently seeks a downstream uridine, or product, to verify selection of the correct cytidine.

RECENT PUBLICATIONS:

Carter CW Jr. Whence the genetic code? Thawing the 'frozen accident'. Heredity. 2008 Apr;100(4):339-40. Epub 2008 Feb 13.

Weinreb V, Carter CW Jr. Mg2+-free Bacillus stearothermophilus tryptophanyl-tRNA synthetase retains a major fraction of the overall rate enhancement for tryptophan activation. J Am Chem Soc. 2008 Jan 30;130(4):1488-94.

Kapustina M, Weinreb V, Li L, Kuhlman B, Carter CW Jr. A conformational transition state accompanies tryptophan activation by B. stearothermophilus tryptophanyl-tRNA synthetase. Structure. 2007 Oct;15(10):1272-84.

Retailleau P, Weinreb V, Hu M, Carter CW Jr. Crystal structure of tryptophanyl-tRNA synthetase complexed with adenosine-5' tetraphosphate: evidence for distributed use of catalytic binding energy in amino acid activation by class I aminoacyl-tRNA synthetases.  J Mol Biol. 2007 May 25;369(1):108-28.

Pham Y, Li L, Kim A, Erdogan O, Weinreb V, Butterfoss GL, Kuhlman B, Carter CW Jr. A minimal TrpRS catalytic domain supports sense/antisense ancestry of class I and II aminoacyl-tRNA synthetases. Mol Cell. 2007 Mar 23;25(6):851-62.

Carter CW Jr, Riès-Kautt M. Improving marginal crystals. Methods Mol Biol. 2007;363:153-74.

Kapustina M, Carter CW Jr. Computational studies of tryptophanyl-tRNA synthetase: activation of ATP by induced-fit. J Mol Biol. 2006 Oct 6;362(5):1159-80.

Roach J, Sharma S, Kapustina M, Carter CW Jr. Structure alignment via Delaunay tetrahedralization. Proteins. 2005 Jul 1;60(1):66-81.