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Characterization of alkaline transitions in ferricytochrome c using carbon-deuterium infrared probes

P. Weinkam, J. Zimmermann, L.B. Sagle, S. Matsuda, P.E. Dawson, P.G. Wolynes, F.E. Romesberg, Biochemistry (2008) 47:13470-13480.
pubpic2008weinkamWe use a combination of experiments and simulations to provide a high-resolution view of the structural changes in cyt c with increasing alkaline conditions. Alkaline-induced transitions were characterized under equilibrium conditions by monitoring IR absorptions of carbon−deuterium chromophores incorporated at Leu68, Lys72, Lys73, Lys79, and Met80. The data suggest that at least four intermediates are formed as the pH is increased prior to complete unfolding of the protein.

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Unnatural substrate repertoire of A, B, and X family DNA polymerases

G.T. Hwang, F.E. Romesberg, J. Am. Chem. Soc. (2008) 130:14872-14882.
pubpic2008hwangWe examine the ability of five different polymerases (family A polymerases from bacteriophage T7 and Thermus aquaticus, family B polymerases from Thermococcus litoralis and Thermococcus 9°N-7, and the family X polymerase, human polymerase β) to replicate DNA containing four different unnatural nucleotides bearing predominantly hydrophobic nucleobase analogs, and we identify some aspects of recognition that are conserved.

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Optimization of the pyridyl nucleobase scaffold for polymerase recognition and unnatural base pair replication

Y. Hari, G.T. Hwang, A.M. Leconte, N. Joubert, M. Hocek, F.E. Romesberg, ChemBioChem, (2008) 9:2796-2799.
pubpic2008hariWe demonstrate that the pyridyl nucleobase scaffold can be optimized for replication, both as a self pair and as a component of a heteropair, and we identify a pyridyl-nucleotide self pair that is well recognized by a DNA polymerase, allowing it to be replicated and used to synthesize site-specifically labeled DNA in good yields.

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Exploring the energy landscape of antibody-antigen complexes: protein dynamics, flexibility, and molecular recognition

M.C. Thielges, J. Zimmermann, W. Yu, M. Oda, F.E. Romesberg, Biochemistry (2008) 47:7237-7247. [June 2008 Hot Article].
pubpic2008bthielgesWe examine a panel of antibodies elicited to the chromophoric antigen fluorescein. On the basis of isothermal titration calorimetry, we select six antibodies that bind fluorescein with diverse binding entropies, suggestive of varying contributions of dynamics to molecular recognition. We find that more than half of all the somatic mutations among the six antibodies are located in positions unlikely to contact fluorescein directly, and using 3PEPS and TG spectroscopy, we find a high level of dynamic diversity among the antibodies.

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Carbon-Deuterium bonds as probes of dihydrofolate reductase

M.C. Thielges, D.A. Case, F.E. Romesberg, J. Am. Chem. Soc. (2008) 130:6597-6603.
pubpic2008thielgesWe report a preliminary analysis of (methyl-d3) methionine residues, Met16, Met20, and Met42, within DHFR. The results confirm the sensitivity of the carbon−deuterium bonds to their local protein environment and demonstrate that dihydrofolate reductase is electrostatically and dynamically heterogeneous.

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Identification of pathways controlling DNA damage induced mutation in Saccharomyces cerevisiae

E.T. Lis, B.M. O’Neill, C. Gil-Lemaignere, J.K. Chin, F.E. Romesberg, DNA Repair (2008) 7:801-810.
pubpic2008lisTo further understand the mutagenic response to DNA damage, we screen a collection of 4848 haploid gene deletion strains of S. cerevisiae for decreased damage-induced mutation of the CAN1 gene, and we identify a pathway of induced mutation that involves a replicative polymerase and that is effectively inhibited by the RNR inhibitor, hydroxyurea.