S. cerevisiae Prp8 and Aar2

Kaylyn Stanton '17 and Emily Olson '17


Model View:

I. Introduction

Before mRNA can be fully synthesized, splicing by the Spliceosome must occur to remove introns from pre-mRNA. This molecule consists of five small nuclear ribonucleoproteins (snRNPs) and multiple protein complexes. Prp8 is one of these protein complexes, located in the U5 snRNP (Grainger et al 2005). The component is highly conserved across most species, with over 60% conservation. It is also the largest component in the spliceosome containing about 2400 amino acids and varying between 230-250 kDa (Dlakic et al 2011). The Prp8 that we will discuss is from Saccharomyces cerevisiae, commonly known as yeast. Together with Brr2 and Snu114, Prp8 plays an important role in spliceosome activation as it is located at the catalytic center of the spliceosome.

Another component of the U5 snRNP is the protein Aar2. This protein is required for the splicing of U3 precursors. Aar2 interacts with both the RNaseH-like domain and the Jab1/MPN domain. These interactions are the basis of the catalytic center of the spliceosome. Aar2 also acts to regulate spliceosome activation by blocking the U4/U6 snRNA binding to Prp8 and stalling spliceosome activation. Understanding the interactions between Aar2 and Prp8 are essential to understand the structure and function of the entire spliceosome.

II. General Structure

Prp8 is comprised of , however the first component, the N-terminal Domain, is not shown. The six other domains are: 1. Reverse Transcriptase (RT) Finger/Palm Domain, 2. RT Thumb/X Domain, 3. Linker, 4. Endonuclease Domain, 5. RNaseH-like, and 6. Jab1/MPN Domain. Each of these domains are detailed in the sections below.


is a smaller protein, containing 355 residues. It can be broken up into the N-terminal Domain and the C-terminal Domain. The C-terminal Domain is the region that interacts with Prp8.

Both the RNaseH-like domain and the Jab1/MPN domain interact with Aar2 by folding in to create a new, larger domain. This domain can be broken down into the and the . These two domains interact through the .

III. Large Polymerase-Like Domain

The large polymerase-like domain is made up of the reverse transcriptase (RT) finger/palm region and the RT Thumb/X region. In the , the palm sub-domain forms a four-stranded antiparallel beta-sheet, flanked by alpha-helices. The main motif in this domain is , found in the loop between RTB7 and 8. Within this motif, there are three aspartates that are incapable of binding divalent metal ions, leading researchers to conclude that it has reverse transcriptase activity. The RT Thumb/X region is made of four anti-parallel beta-sheets (RTB10-RTB13) and a 3-alpha-helix bundle (RTa14-RTa16) (Galej et al, 2013).

IV. Small Type II Restriction Endonuclease-Like Domain

The of Prp8 comprises the small type II restriction endonuclease-like domain. It is made up of five beta-strands: EnB1, EnB2, EnB4, EnB5, and EnB6; and three alpha-helices: Ena1, Ena2, and Ena3. This domain forms a cavity along with RNaseH-Like and Thumb domains that holds the catalytic core of the group II intron RNA. While it is structurally similar to an endonuclease domain of the influenza virus (which has catalytic activity), mutations in the conserved elements of the endonuclease-like fold does not affect the molecule. This suggests that this region is conserved for structural and stability reasons (Galej et al, 2013).

IV. Prp8 and Aar2 Interaction

The component directly interacts with the Aar2 protein through its RNaseH-like domain and Jab1/MPN domain. The C-terminal of the Aar2 protein forms a beta-sheet. This sheet together the beta-hairpin of RNase-like domain and the beta-barrel of the Jab1/MPN domain.

The two proteins also interact through two alpha-helices of the RNaseH-like region of Prp8 and the C-terminal helical domain of Aar2. An arginine located at the tip of the RT-finger region interacts with an asparagine and a leucine in the helical region of Aar2. Other contacts between Prp8 and Aar2 occur through the linker. occur between Aspartic Acid, Glycine, and Glutamic Acid of Aar2 and Arginine, Glutamine, and another Arginine of the Prp8 linker, respectively (Galej et al 2011).

VI. References

Dlakic, M., and A. Mushegian. "Prp8, the Pivotal Protein of the Spliceosomal Catalytic Center, Evolved from a Retroelement-encoded Reverse Transcriptase." RNA 17.5 (2011): 799-808. PMC. Web.

Galej, Wojciech P., Chris Oubridge, Andrew J. Newman, and Kiyoshi Nagai. "Crystal Structure of Prp8 Reveals Active Site Cavity of the Spliceosome." Nature 493.7434 (2013): 638-43. PMC. Web.

Grainger, Richard J., and Jean Beggs. "Prp8 Protein: At the Heart of the Spliceosome." RNA 11.5 (2005): 533-57. PMC. Web.

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