View source for Prp40

<StructureSection load='1o6w' size='450' side='right' scene='Sandbox_504/Start_scene/1' caption=''>
== Structure of Prp40 ==
by Kelly Hrywkiw

__TOC__
 
=Introduction=

'''Prp40''' has been implicated in early steps of [http://en.wikipedia.org/wiki/Spliceosome spliceosomal] assembly, and binding to the phosphorylated C-terminal domain (phospho-CTD) of [http://en.wikipedia.org/wiki/RNA_polymerase_II RNA polymerase II] (RNAPII)<ref name ="bonet">PMID:19722265</ref>.  Two types of domains are found in Prp40, two [http://en.wikipedia.org/wiki/WW_domain WW] and six consecutive FF domains<ref name ="wiesner">PMID:12460579</ref>.    WW domains contain two highly conserved tyrosine residues that bind to proline rich sequences, thereby mediating protein-protein interactions<ref name ="wiesner"/>. Both WW domains of Prp40 can interact with PPxY motifs (x is any residue) and PPѰѰP motifs (Ѱ is an aliphatic residue)<ref name ="wiesner"/>.  FF domains contain two highly conserved proline residues and can be found in arrays up to six domains<ref name ="gasch">PMID:16253993</ref>.  Interestingly, these tandem domains appear not to function in co-operative binding, rather they may provide multiple independent binding sites<ref name ="gasch"/>.  With regards to interactions between the different domains, the WW domains bind to the splicing protein: branch point-binding protein (BBP, also known as Ms15 and ySF1) and Prp8; the FF domains interact with the splicing factor Clf1 (Syf3p in yeast), and the phospho-CTD of RNAPII<ref name ="gasch"/><ref name ="wiesner"/>.

=Role in spliceosomal assembly=

The processing of [http://en.wikipedia.org/wiki/Pre-mRNA pre-mRNA] takes place through the use of a large dynamic machine known as the spliceosome, through which [http://en.wikipedia.org/wiki/Intron introns] are removed and [http://en.wikipedia.org/wiki/Exon exons] are spliced together to create a mature [http://en.wikipedia.org/wiki/MRNA mRNA]<ref name ="architecture of"/><ref name ="structure and function">PMID:19000813</ref>.  The spliceosome is comprised of five [http://en.wikipedia.org/wiki/SnRNA snRNA] molecules (snRNAs U1, U2, U4, U5, and U6) and over one hundred associated proteins<ref name ="architecture of"/><ref name ="structure and function"/>.  Assembly of the spliceosome is thought to take place in a stepwise manner around the pre-mRNA transcript<ref name ="architecture of"/><ref name ="structure and function"/>.  The first step involves recognition of the 5’ splice site (ss) by U1 snRNP, followed by recognition of the branch point sequence by U2 snRNP<ref name ="architecture of"/><ref name ="structure and function"/>.  From this point the tri-snNRP consisting of U4/U6•U5 binds<ref name ="architecture of"/><ref name ="structure and function"/>.  Together the five snRNPs form the precatalytic spliceosome which must undergo a series of changes before it can actively splice<ref name ="structural evidence">PMID:19525970</ref><ref name ="structure and function"/><ref name ="common design">PMID:19525970</ref><ref name ="architecture of">PMID:22471593</ref>.

Prp40 participates in cross-intron bridging through interactions with the 5’ss and the BBP, which brings the 5’ss and the branch point into spatial proximity<ref name ="wiesner"/>.  In addition, Prp40 interacts with Prp8, such that it is possible that while one W domain interacts with Prp8 the other interacts with the 5’ss at the same time<ref name ="wiesner"/>.  In fact, this interaction is thought to bridge the 5’ss to the 3’ss after U2 snRNP association displaces BBP and causes Prp8 to interact with the tri-snRNP at the 3’ss.   

=Role in transcription=

RNAPII is responsible for mRNA synthesis, the first step of gene expression of eukaryotes<ref name ="cramer">PMID:11313498</ref>.  During the synthesis cycle of mRNA, RNAPII interacts with up to six general [http://en.wikipedia.org/wiki/Transcription_factor transcription factors] and numerous regulatory proteins.  It is comprised of 12 subunits (Rpb1 through 12) and several disordered regions including the NH2 tail of Rpb6 and Rpb12, short exposed loops in Rpb1, 2, and 8, and the CTD of the largest subunit Rpb1<ref name ="cramer"/>.  The CTD is subject to hyperphosphorylation, such that RNAPII can exist in either a hyperphosphorylated form (RNAPII0) or a non-phosphorylated form (RNAPIIA) at the CTD<ref name ="phatnani">PMID:17079683</ref>.  The phospho-CTD helps coordinate pre-mRNA processing events such as localizing and activating the 5’capping complex.  Interestingly, when RNAPII0 is transcriptionally active in the nuclear matrix it colocalizes with splicing factors<ref name ="morris">PMID:10978320</ref>. 
 
Prp40 can bind to the RNAPII0 at the phospho-CTD<ref name ="wiesner"/><ref name ="morris"/>.  The WW domains of Prp40 have been shown to interact with the phospho-CTD; however, they do not interact with the conserved YSPTSPS sequence of phospho-CTD.  The FF domains have been shown to interact with the phosphor-CTD repeats<ref name ="gasch"/>.  Not all the FF domains of Prp40 exhibit the same functionality, such that the first FF domain and the fourth FF domain do not interact with the phosphor-CTD<ref name ="gasch"/>.  The ability of Prp40 to bind to spliceosomal factors and the phosphor-CTD of RNAPII0 could connect the phosphor-CTD to the earliest stages of spliceosomal commitment complex formation<ref name ="morris"/>. 
 
=Structure of Prp40=

Yeast Prp40 contains 583 residues, two WW domains, and four FF domains connected through amino acid linkers<ref name ="morris"/>. 

[[Image:Prp40 domains.PNG|thumb|left|upright=4|alt=Proposed domains.|Figure1: Schematic representation of the domain organization in Prp40.]]  

==The WW domains==


<Structure load='1o6w' size='250' thumb='false' align='right' caption='Figure 2: N terminal (blue) to C terminal (red) ribbon representation of the FF1 domain of Prp40, [[1o6w]]' scene='Sandbox_504/Start_scene/1'/>
 

The overall structure of the two consecutive WW domains follows that of one triple curved antiparallel β-strand sheet connected to the other by an α-helical linker composed of residues <scene name='Sandbox_504/Linker/1'>Lys30-Glu42</scene>. The residues in each of the β strands are as follows, β1 <scene name='Sandbox_504/Wwdomain1_b1/2'>(Trp4-Asp9)</scene>, β2 <scene name='Sandbox_504/Wwdomain1_b2/1'>(Arg13-Thr19)</scene>, and β3 <scene name='Sandbox_504/Wwdomain1_b3/1'>(Lys22-Trp26)</scene> in the <scene name='Sandbox_504/Ww1/2'>first W domain</scene>, and β1 <scene name='Sandbox_504/B1_of_ww2/1'>(Trp45-Thr50)</scene>, β2 <scene name='Sandbox_504/B2_of_ww2/1'>(Lys54-Pro60)</scene>, and β3 <scene name='Sandbox_504/B3_of_ww2/1'>(Arg63-Trp67)</scene> in the <scene name='Sandbox_504/Ww2/1'>second W domain</scene>.  Located on the convex surface of each of the domains lie three residues, <scene name='Sandbox_504/Hydrophobicresiduesandprotein/1'>Trp4/45, Tyr16/57, and Pro29/Leu40</scene>, that form a <scene name='Sandbox_504/Hydrophobicresiduesandprotein/2'>hydrophobic core</scene>.  On the concave surface lies an aromatic pocket comprised of the residues <scene name='Sandbox_504/Aromatic_pocket_residues/1'>Tyr15/56, Tyr17/58, and Trp26/67</scene>.  These pockets makes up the ligand binding sites on each of the WW domains, however they do not form one large pocket, rather <scene name='Sandbox_504/Aromatic_pocket_residues/2'>two separate small pockets </scene> that face away from each other.  The linker residues Leu32 and Leu40 fold back into the hydrophobic cores of the WW domains<ref name ="wiesner"/>. 

==The first FF domain (FF1)==


<Structure load='1o6w' size='250' thumb='false' align='left' caption='Figure 2: N terminal (blue) to C terminal (red) ribbon representation of the consecutive WW domains of Prp40, [[1o6w]]' scene='Sandbox_504/Start_scene_ff1/1'/>

The FF1 domain of Prp40 is comprised of three alpha helices, and one 310 helix located between α2 and α3.  Helices are composed of the following residues, <scene name='Sandbox_504/Ff1_a1/1'>α1</scene> (134-146), <scene name='Sandbox_504/Ff1_a2/2'>α2</scene> (154-163), <scene name='Sandbox_504/Ff1_310/1'>310</scene> (167-170), and <scene name='Sandbox_504/Ff1_a3/1'>α3</scene> (175-187).  The core domain is made up of a series of aromatic and aliphatic residues.  A type 1 β-turn is exhibited by the residues Asp149, Ser150, Thr151, and Trp152<ref name ="gasch"/>.

==The fourth FF domain (FF4)==


<Structure load='1o6w' size='250' thumb='false' align='right' caption='Figure 2: N terminal (blue) to C terminal (red) ribbon representation of the consecutive WW domains of Prp40, [[1o6w]]' scene='Sandbox_504/Start_scene_ff4/1'/>

Domain FF4 of Prp40 exhibits compact four helical bundle fold comprised of an α1-α2-310-α3 topology.  The composition of each helix is a follows <scene name='Sandbox_504/A1_of_ff4/1'>α1</scene> (Glu489-Thr507), <scene name='Sandbox_504/A2_of_ff4/1'>α2</scene> (Trp519-Leu526), <scene name='Sandbox_504/310_of_ff4/1'>310</scene> (Tyr532-Gly536) and <scene name='Sandbox_504/A3_of_ff4/1'>α3</scene> (Asp539-Phe549). There are a series of interactions between the different helices, for example Tyr532 is in contact with Phe500, Leu503, Ser523, and Arg542.  A difference between FF1 and FF4 is the presence of five extra amino acids in F4 which gives the <scene name='Sandbox_504/Loop_of_ff4/1'>loop</scene> located between α 1 and α2 an extra turn.  This insertion however does not increase the flexibility of F4 as compared to F1<ref name ="bonet"/>.   

=Additional Resources=
*[http://www.rcsb.org/pdb/explore/explore.do?structureId=2KFD Prp40 FF4 domain, in the RCSB Protein Data Bank] 
*[http://www.rcsb.org/pdb/explore/explore.do?structureId=1O6W SOLUTION STRUCTURE OF THE PRP40 WW DOMAIN PAIR OF THE YEAST SPLICING FACTOR PRP40, in the RCSB Protein Data Bank]
*[http://www.rcsb.org/pdb/explore/explore.do?structureId=2B7E First FF domain of Prp40 Yeast Protein, in the RCSB Protein Data Bank] 


=References=
<References/>