Lester Floyd Harris, Principal Investigator
Leonard S. Schultz, Co-Principal Investigator

Dynamic Simulations of Solvated Protein/DNA Complexes

Computer models of the mouse mammary tumor virus (MMTV) nucleosome structure and its components: a) the histone core is made up of an octamer composed of two each of histone proteins H2A, H2B, H3, and H4; b) 146 base pair MMTV LTR DNA wrapped in a superhelical conformation; c) the superhelical DNA wrapped around the histone protein octamer core; d) the MMTV nucleosome with five glucocorticoid receptor proteins docked at their response element DNA sequences.

These researchers conducted experiments investigating the mechanism(s) of a genetic switch controlled by deoxyribonucleic acid (DNA) regulatory proteins. Interest lay in steroid hormone receptor protein interaction with DNA in the pathogenesis of breast cancer. The researchers previously reported on a mechanism describing how these DNA regulatory proteins recognize and bind to their specific sites on DNA. In the most recent research period, they conducted molecular dynamics simulations in solvent to investigate hydrogen bonding, van der Waals and electrostatic interactions between amino acids of the DNA regulatory proteins, and nucleotides of their cognate DNA binding sites. They used a biological model of a virus that infects bacteria, a bacteriophage, for investigating the genetic switch controlling lytic/lysogeny expression in the 434 bacteriophage. It is known that this genetic switch is under the control of two proteins of the bacteriophage, cI repressor and Cro, that interact with DNA sites, operators OR1, OR2, and OR3, within the bacteriophage genome. The cI repressor protein preferentially binds first to OR1 then to OR2 inducing transcription to the left. In contrast, the Cro protein preferentially binds first to OR3 then OR2 inducing transcription to the right. The basic molecular interactions between amino acids from the bacteriophage proteins interacting with specific regulatory sites on DNA are in agreement with the researchers’ earlier findings for the amino acids of the steroid hormone receptor proteins interacting with their regulatory sites on DNA. Together these findings strongly support their theory that proteins recognize specific sites on DNA based on stereochemical complementarity and conservation of genetic information between the proteins and the specific sites on DNA to which they bind and turn genes off or on.

While bacteriophage genome, lacking a nucleus and chromosomes, is very simple in its organization and structure, the genome of higher forms of life is very organized, compact, and structurally complex, and contains a nucleus and chromosomes. The DNA of the chromosomes is packaged in a complex structure, chromatin, with the aid of specialized proteins called histones. The fundamental packing unit of the chromatin is the nucleosome, which is conserved, in all higher forms of life. The nucleosome consists of 146 nucleotide base pairs wrapped around a histone core (see figure, parts a–c). The histone core is made up of an octamer composed of two each of histones: H2A, H2B, H3, and H4 (see part a of figure). Recently, a nucleosome structure has been crystallized and its atomic co-ordinates have been determined by x-ray crystallography. These researchers used these atomic coordinates as a template to construct a nucleosome model of the 146 base pair DNA sequence of the long terminal repeat, LTR, flanking the mouse mammary tumor virus genome that is rich in binding sites for steroid hormone receptor proteins. This LTR DNA sequence contains five specific DNA binding sites, glucocorticoil response elements (GREs), for the glucocorticoid receptor protein that is involved in genetic regulation of the mouse mammary tumor virus genome. These researchers use the mouse mammary tumor virus as a model for studying steroid hormone regulation of genes in breast cancer. As seen with the bacteriophage operator DNA sequence of the genetic switch, the GREs within the LTR DNA sequence function in concert to regulate the mouse mammary tumor virus genome. Earlier, these researchers conducted molecular dynamics simulations on one of the five GREs of the LTR as an extracted DNA sequence containing a GRE and its flanking nucleotide region in complex with the glucocorticoid receptor protein. They were able to characterize the atomic interactions between the protein and the DNA identifying binding structures and amino acids on the protein and specific nucleotides within the GRE and its flanking region. They are now beginning molecular dynamics simulations on the LTR nucleosome model with multiple glucocorticoid receptor proteins docked onto the five GRE sites (see part d of figure). This nucleosome model will allow the researchers to study eukaryotic regulatory elements in their natural context to develop a better understanding of the requirements for controlling expression of transfected genes.

Research Group

Pamela D. Popken-Harris, Research Associate
Michael R. Sullivan, Research Associate