Dr. Jim Nolan is a molecular biologist studying ribozyme structure-function and RNA-protein interactions. He also is using bioinformatic approaches to study viral genome evolution and bioprocess development.

Research Description

We are examining structural aspects of RNA-protein interactions using the bacterial ribonuclease P (RNase P) holoenzyme as a model system. RNase P consists of a large RNA and a small protein subunit, which together cleave 5'-precursor sequences from tRNA transcripts. In vitro at high ionic strength, the RNA subunit is the catalytic moiety and the protein is dispensable for catalysis. Although the RNase P protein is not required for activity in vitro, it is essential for viability in vivo. In addition, the kinetics of the holoenzyme reaction in vitro differs dramatically from the RNA-alone reaction in salt requirement, substrate specificity, and reaction rate.

We are using a detailed structural analysis of the RNase P RNA-protein complex to investigate the how the protein binds to RNase P RNA, and how this binding affects the reaction mechanism, including changes in how the RNase P RNA interacts with its tRNA substrate in the presence of RNase P protein. These problems are being studied by a variety of techniques. We have incorporated crosslinking reagents at specific sites of both the RNA and protein subunits to further characterize the sites of interaction between the two subunits. Regions of the RNase P protein important for interaction with RNase P RNA were identified by in vitro and in vivo analyses of mutations in the RNase P protein.

RNase P protein has an unusual protein-folding motif, a left-handed crossover connection as part of central sheet RNA binding cleft. This unusual structure is shared with ribosomal protein S5 and elongation factor EF-G. Together, the proteins represent a novel class of RNA binding protein although they lack significant sequence homology between the group members. This structural conservation among widely divergent sequences will be exploited to investigate important determinants of RNase P protein function as well as determinants responsible for folding of the protein domain.

In collaboration with Jim D. Karam, we are performing a genomic sequence analysis of T4-like bacteriophages to survey global evolution of viral genomes. The genome sequences will provide phylogenetic variant sequences of known proteins and will validate predicted small open reading frames (ORFs). Results will be disseminated via the Phage Genome Browser (phage.bioc.tulane.edu), which highlights genome features and provides links to gene-specific information.

In collaboration with Joan Bennett, we are investigating fungal genomics for biomass conversion. We seek to identify new protein sequences from databases using bioinformatics tools to expand capabilities for fungal bioprocess development.

We are also engaged in collaborative efforts with Joseph Lasky to use ribozymes to inactivate cytokine mRNAs in mouse.