The long term objective of this proposed work is to develop new strategies to allow the systematic study of the relationship between the structure and function of proteins. The principle new approaches involve creating specific alterations in the amino acid sequence of a protein by introducing corresponding changes in the structural gene encoding that protein. These approaches will be first applied to enzyme Beta-lactamase, which is responsible for the resistance to penicillin therapy of an ever-increasing number of strains of infectious microorganisms. The principle methodologies employed include: chemical synthesis of deoxyoligonucleotides; deoxyoligonucleotide-directed, site-specific mutagenesis; random mutagenesis; biophysical studies of the structural and functional characteristics of the resulting mutant proteins; nmr observations both of mutant proteins and of hybrids of synthetic deoxyoligonucleotides of the types involved in the mutagenesis techniques. To understand the mechanism of action of Beta-lactamase should be of great importance in allowing the design of new antibiotics of the penicillin/cephalosporin type that are not hydrolyzed and inactivated by the enzyme. This should provide powerful new agents to combat infections by the many strains of microorganisms that are presently resistant to many penicillin-type antibiotics. To be able to create and study many structural analogues of a parent protein will provide striking new insights into how proteins carry out their myriad functions (as, for example, biological catalysts, hormones, transport agents, cell surface receptors, structural elements in cells and organisms, muscles that convert chemical energy into work, antibodies that distinguish self from non-self); it will also give us eventually the ability to design proteins with specific, novel and useful properties.
| Fernandez, C O; Cricco, J A; Slutter, C E et al. (2001) Axial ligand modulation of the electronic structures of binuclear copper sites: analysis of paramagnetic 1H NMR spectra of Met160Gln Cu(A). J Am Chem Soc 123:11678-85 |
