Multi-Functional Activities of a Group I Intron-encoded Protein

Temple University, Philadelphia PA

Investigators

Abstract

The objective of this project is to understand the relationship between two activities of a protein encoded by a group I intron. The protein facilitates a splicing reaction that excises the intron from its precursor RNA and also acts as a DNA endonuclease, cleaving DNA at a specific sequence. It is commonly believed that the RNA splicing activity evolved from a protein that originally functioned solely as a DNA endonuclease. However it is unlikely that this new function arose simply by fusion of an additional gene sequence. Preliminary evidence indicates that the DNA and RNA substrates bind to distinct or partially overlapping sites on the protein. The question then arises as to how a protein acquires a new function and how easily this can be detected. This is particularly relevant to annotating genomes. Attributing a single function to a gene may constitute only partial characterization of the sequence since an unknown number of proteins may "moonlight" and perform a second, unrelated and unforeseen task. Group I introns can catalyze their own excision from RNA precursors. An intron-encoded protein, called a maturase, significantly facilitates removal of such an intron in Aspergillus nidulans. It also has DNA endonuclease activity. The three-dimensional structure of the protein has been obtained with the help of a collaborator and the region which binds the cleavable DNA sequence has been identified. Guided by this model, the relationship between the binding sites of the RNA and DNA substrates will be studied biochemically and genetically. Current diffraction data from crystals of group I introns are of insufficient quality to reveal structural details at the atomic level. Co-crystallizing RNA with a protein can often solve this problem. A collaboration will be performed to obtain a high-resolution structure of the intron-maturase complex. This will not only reveal the RNA binding site on the protein and help to determine how the RNA binding site arose, but also provide a wealth of structural information about the group I intron RNA itself.