News and Publications

Computer Modeling of Proteins and Nucleic Acids

D.A. Case, M. Crowley, T. Dwyer,* V. Feher, F. Himo, H. Liu, T. Liu, T. Macke, G. Mer, A. Onufriev, R. Torres, V. Tsui, M. Ullmann, X. Xu

* University of San Diego, San Diego, CA

Computer simulations offer an exciting approach to the study of many aspects of biochemical interactions. In our group, we focus primarily on molecular dynamics simulations (in which Newton's equations of motions are solved numerically) to model the solution behavior of biomacromolecules. Recent applications include detailed analyses of electrostatic interactions in short peptides (folded and unfolded), proteins, and oligonucleotides in solution.

In addition, molecular dynamics methods are useful in refining solution structures of proteins by using constraints derived from nuclear magnetic resonance spectroscopy, and we continue to explore new methods in this area. Our developments in these areas are incorporated into the Amber molecular modeling package, designed for large-scale biomolecular simulations, and into other software, including Nucleic Acid Builder, for developing 3-dimensional models of unusual nucleic acid structures; SPASM, for analyzing chemical shifts in proteins and nucleic acids; and RNAMotif, for finding structural motifs in databases of genomic sequences.

PROTEIN AND NUCLEIC ACID STRUCTURE AND DYNAMICS

Our overall goal is to extract the maximum amount of information on biomolecular structure and dynamics from nuclear magnetic resonance experiments. To this end, we are studying the application of direct refinement methods for using nuclear magnetic resonance data to determine biomolecular structures in solution, going beyond distance constraints to generate closer connections between calculated and observed spectra. We are also using quantum chemistry to study chemical shifts and spin-spin coupling constants. New types of information, such as chemical shift anisotropies, direct dipolar couplings in partially oriented samples, and analysis of cross-correlated relaxation, are also being used to guide structure refinement. Recent structural studies focused on transcription factors and minor groove-binding drugs complexed with DNA.

NUCLEIC ACID MODELING

Another project centers on the development of novel computer methods to construct models of "unusual" nucleic acids that go beyond traditional helical motifs. We are using these methods to study circular DNA, small RNA fragments, and 3- and 4-stranded DNA complexes, including models for recombination sites. We continue to develop efficient computer implementations of continuum solvent methods to allow simplified simulations that do not require a detailed description of the solvent (water) molecules; this approach also provides a useful way to study salt effects.

This research is part of a larger effort to develop "low resolution" models of nucleic acids that can be extended to much larger structures such as circular DNA, viruses, or models of ribosomal particles. A computer language, NAB, has been developed to make it easier to construct initial molecular models for complex and often low-resolution problems. The language is being used to construct models for the small subunit of the ribosome and for the analysis of curved and circular DNA.

DYNAMICS AND ENERGETICS OF NATIVE AND NONNATIVE STATES OF PROTEINS

Analysis methods similar to those described for nucleic acids are being used to estimate thermodynamic properties of "molten globules" and unfolded states of proteins. These studies are an extension of our earlier work on the folding of peptide fragments of proteins. A key feature is the development of computational methods that can be used to model pH and salt dependence of complex conformational transitions, such as unfolding events.

A second goal of this work is a detailed interpretation of nuclear magnetic resonance results for protein nonnative states through molecular dynamics simulations and the construction of models for molecular motion and disorder. In a parallel effort, we are studying correlated fluctuations about native conformations in dihydrofolate reductase and in metallo-ß-lactamase in an effort to make more secure connections between the motions of proteins and the activities of enzymes.

SEQUENCE ANALYSIS OF RNA

We developed a software program, RNAMotif, that is used to search a database for RNA sequences that match a motif associated with secondary structure interactions. A match means that the given sequence can adopt the given secondary structure, and matches can be ranked by applying scoring rules that may provide finer distinctions than simply matching to a profile.

We find that it is useful to combine results from motif searches with estimates of folding energetics, because the latter are effective in eliminating false-positives. We have had good preliminary success in identifying structural RNA motifs in genome sequences, including tRNA genes, iron-response elements, and transcriptionally active pseudoknots.

Figure 1 illustrates how a search might be done for tRNA genes in the yeast Saccharomyces cerevisiae genome. The program searches for a fairly generic cloverleaf secondary structure (top), requiring also that a few key conserved nucleotides be present. This step finds 272 of 275 currently known tRNA genes, along with many false-positives (solutions that match the profile but are probably not tRNAs). Ranking the matches according to their predicted folding free energies (bottom) provides a simple way to distinguish true from false hits. This example shows how novel descriptors for RNAs with interesting structural characteristics might be generated and optimized; we plan extensions to other RNA structural motifs.

PUBLICATIONS
Case, D.A., Scheurer C., Brüschweiler, R. Static and dynamic effects on vicinal scalar J couplings in proteins and peptides: A MD/DFT analysis. J. Am. Chem. Soc. 122:10390, 2000.

Cornell, W., Abseher, R., Nilges, M., Case, D.A. Continuum solvent molecular dynamics study of flexibility in interleukin-8. J. Mol. Graph. Model. 19:136, 2001.

Kollman, P., Massova, I., Reyes, C., Kuhn, B., Huo, S., Chong, L., Lee, M., Lee, T., Duan, Y., Wang, W., Donini, O., Cieplak, P., Srinivasan, J., Case, D.A., Cheatham, T.E. III. Calculating structures and free energies of complex molecules: Combining molecular mechanics and continuum models. Acc. Chem. Res. 33:889, 2000.

Ogawa, A.K., Abou-Zied, O.K., Tsui, V., Jimenez, R., Case, D.A., Romesberg, F.E. A photo-tautomerizable model DNA base pair. J. Am. Chem. Soc. 122:9917, 2000.

Onufriev, A., Case, D.A., Ullmann, G.M. A novel view of pH titration in biomolecules. Biochemistry 40:3413, 2001.

Smith, J.A., Bifulco, G., Case, D.A., Boger, D.L., Gomez-Paloma, L., Chazin, W.J. The structural basis for in situ activation of DNA alkylation by duocarmycin SA. J. Mol. Biol. 300:1195, 2000.

Sosa, C.P., Hewitt, T., Lee, M.S., Case, D.A. Vectorization of the generalized Born model for molecular dynamics on shared-memory computers. J. Mol. Struct. Theochem, in press.

Tsui, V., Radhakrishnan, I., Wright, P.E., Case, D.A. NMR and molecular dynamics studies of the hydration of a zinc finger-DNA complex. J. Mol. Biol. 302:1101, 2000.

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