Norman Pace - Professor

Our laboratory carries out research in two areas. In one, we study the structure and mechanism of activity of the catalytic RNA moiety of the RNA processing enzyme ribo-nuclease P (Rnase P). In the second, we explore naturally occurring communities of microorganisms using molecular methods that sidestep the need to cultivate the organisms. Although these two research areas might seem disparate, they overlap substantially in techniques and in a phylogenetic-comparative approach.

Rnase P forms one mature end of all tRNAs by cleaving off a precursor RNA sequence. Rnase P consists of both protein and RNA (ca. 400 nt) subunits; the RNA is the catalyst. In order to understand the chemistry and biology of Rnase P RNA, we need to know its structure. Over the past several years we have worked out the secondary structure of this large RNA and learned much about its mechanism. Patterns of conservation of sequence and structure are used as guides for experiments involving mutagenic and kinetic analysis to study the mechanisms of substrate binding and catalysis by Rnase P. Current efforts focus on structure as it relates to the mechanism of Rnase P, and on crystallization of Rnase P components for atomic-level structure determinations.

In the second project area, comparisons of ribosomal RNA (rRNA) sequences are used to explore naturally occurring microbial ecosystems. Little is known about the natural microbial world because biologists have always had to cultivate organisms in order to study them, and typically <0.1% of organisms observed microscopically in the environment can be cultivated. Consequently, we have been developing a suite of molecular techniques, based on the molecular phylogeny of 16S rRNA, that allow us to characterize naturally occurring organisms without cultivation. Comparisons of 16S rRNA sequences can be used to generate evolutionary trees that can be used as a quantitative framework in which to identify unknown organisms. In natural population analyses, rRNA genes are obtained by cloning from DNA isolated directly from the environment. Sequence and phylogenetic analyses of the rRNA genes incisively relate unknown organisms to known ones. Some properties of the unknown organisms can be inferred from the properties of their relatives. The sequences can be used to design hybridization probes for further studies of interesting organisms, in the environment or laboratory. We have developed these techniques in the context of studying several types of ecosystems. Some examples are sulfide-oxidizing and other symbioses between bacteria and animals, high-temperature ecosystems (Yellowstone and submarine hydrothermal vent environments), hydrocarbon-damaged aquifers, marine picoplankton, deepsubsurface environments, and medical syndromes such as tuberculosis and Crohn’s Disease. Many new organisms have been discovered, including some profoundly different from known ones.