In this work, three projects concerning the creation and control of catalytic processes are presented. In Chapter 1, a project that resulted in the characterization of an antibody able to catalyze the hydrolysis of two epimeric phosphate triesters is presented. Enzymes are thought to catalyze reactions by stabilizing high-energy transition states. Catalytic antibodies, in turn, have been generated that bind to stable analogs of transition states. The transition state for phosphate triester hydrolysis is pentacoordinate, of trigonal bipyramidal (TBP) structure and negatively charged. Stable compounds containing TBP centers are rare in organinc chemistry. As such, antibodies able to bind charged haptens were screened for their ability to catalyze the hydrolysis of homologous phosphate triesters. One antibody was a particularly efficient catalyst. It is therefore possible that structural elements of the amine-oxide hapten that induced the catalytic antibody may be applicable to the generation of other phosphoryl-transfer catalysts. A scheme for the selection of catalytic metalloantibodies is presented in Chapter 2. The display of libraries of antibody fragments on the surface of phage permitted the selection of antibodies able to coordinate metal ions. Antibody fragments were selected for their ability to coordinate transition metals. Such transition metals are cofactors for many hydrolytic enzymes and are used in many systems to carry out the oxidative cleavage of RNA, DNA and proteins. The metal-binding antibodies were also selected for their ability to bind DNA although no DNA cleavage activity was observed. Finally, a previously unappreciated property of the signals that target organelle-specific proteins to their subcellular sites of action is described in Chapter 3. Such targeting sequences are shown to be polymorphic. A polymorphism was revealed upon cloning and the mitochondrial manganese-containing superoxide dismutase from cell lines of normal individuals and patients with genetic diseases of premature aging. The polymorphism consists of a single nucleotide change in the region of the DNA that encodes the signal sequence such that either an alanine or valine is present. Subsequently, eight cell lines and genomic DNA from nineteen individuals were analyzed. Heterozygotes as well as both homozygotes, all three possible combinations of the two signal sequences, were observed. Such signal sequence polymorphisms could result in diseases of distribution, where essential proteins are not properly targeted, thereby leading to absolute or relative deficiencies of critical enzymes within specific cellular compartments. Some features of aging may be interpreted as resulting from such disease.
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