Ribosomal Protein Gene in Yeast

This web page was produced as an assignment for an undergraduate course at Davidson College.

YDR466W Protein in Saccharomyces cerevisiae

Introduction

The YDR466W gene in Saccharomyces cerevisiae encodes for a protein that is still unknown both in structure and function. However, it is located on chromosome 4, near other important genes (such as MRPL28), and could be very important to the study of gene location in the genome and the relationship that occurs when genes are closely located on the same chromosome.

Protein Information

The YDR466W gene in Saccharomyces cerevisiae codes for an unknown protein with a length of 898 amino acids. This protein has not been crystallized yet, and thus there is not a chime file showing the 3D structure of the protein. When a BLAST search was performed, it was found that the most similar protein is the cAMP-dependent protein kinase catalytic subunit in Rattus norvegicus . Therefore, it could be assumed that this unknown protein has some function in the protein kinase cell pathway in Saccharomyces cerevisiae. An attempt in Swiss-Model to determine similar 3D structures did not produce any documented proteins with a 3D stucture similar to YDR466W with an E-value of <0.0001. A PREDATOR search reveals that the protein is made up of 18.15% alpha-helices, 68.04% random coils, and 13.81% extended strand. A hydropathy plot was then conducted, and it was found that the protein had no region where the protein would exceed the value of statistical significance (2.0) for hydrophobic regions. The amino acid sequence for the YDR466W protein is displayed below:

mtsrkrsphd fifkeelghg systvfkald kkspnkiyai kvcskkhiik eakvkyvtie kntmnllaqk hhagiiklyy tfhdeenlyf vldfapggel lsllhkmgtf ndiwtrhfta qlidalefih shgiihrdlk penvlldrdg rlmitdfgaa atidpslsgd sakfnsdsng skdnqncasf vgtaeyvspe lllynqcgyg sdiwalgcmi yqfvqgqppf rgenelktfe kivaldypwg pnnrinnsts pinplvinlv qkilvievne risleqikrh pyfskvdwnd kikiwrgiwq sqgqslqqtt lglpnipqni lptrqlhvid tparsiqitk qkrkkptkis nttssivvwr krlgistgkd dlgtvpsttp avtapndtnv ltntaahsta nialppnsqs nqvkraqlva pnrippkvpv indnvrnksi prtkpnvppl qtssipqkls tssassalsa psteirnqdl thtldgrnsi dihvlkqdyv fiygipyehe gpamslnsyn kidndlitsl vaqhkeelkn sesflqvltl kksgmlsykn tvmegnddqe nkehqmanie dtdlsmydfe fneltrkgfl ilekyknriw fislpsystl skipfnavks stinnnenwv dcffrarqll eekqildkis nvsfdskass epsspppisr kerplsignn vttlsytakn gsqnnapqnd nvgeekpfri psstkdrpga nstpssrhpr vlssnnaget pkkmngrlpn sapstntytn gsvpafnhrp stnvgnnkhn iltskkqgss vfspsssttk pqikttgyrq ptpspplpqm efpttrekys apsnmvisss ryevlhtlnn sqtnfdreia srgasaafrs lqkskkkk

Databases Searched

PREDATOR (results above)

Swiss-Model (results above)

PSIPRED v2.1 Protein Structure Prediction Server

3Dpssm

Sanger Institute pfam

Scop Superfamily

Protein Structure Initiative

Database of Interacting Proteins

What Is There?

Enzymes and Metabolic Pathways Database

Yale Gerstein Lab

Triples Database

PDF Protein Interaction Files

Database Results

PSIPRED

PSIPRED results show the predicted secondary structure for the protein. The results for the YDR466W protein show that the protein has several regions of "strand" structure - much more than in the YmL28 protein. The YDR466W protein also shows many regions of "coils," and others with "helix" regions. The YDR466W protein seems much more balanced in its percentages of each type of secondary structural fold than YmL28.
The MEMSAT Results show that the YDR466W protein probably does not contain a transmembrane helix. This is an important test to determine the subcellular location of the protein, which would help to design experiments to isolate and identify its true functions.
GenTHREADER Results show that portions of the YDR466W protein show a high resemblance to other PDB proteins. These conserved domains are a great find, something that could assist greatly in the identification of the function and structure of YDR466W. Also, these domains could be reused once YDR466W is crystallized in order to fine tune the algorithms used to determine 2D and 3D protein shape predictions. This only makes our databases that much more accurate. Right now, the highest probability match for the first part of YDR466W is the 1KOB protein, which is a twitchin kinase fragment from Aplysia californica. This initial similarity to a kinase protein will be confirmed throughout the searches for YDR466W.

Prof

Prof did not contain any information about the secondary structure of YDR466W.

3Dpssm

3Dpssm Results show that the YDR466W protein is very similar to known protein kinases. In fact, the YDR466W protein brings up 20 proteins that show similarity, and all of their folds are described as protein kinase-like. The protein that shows the most similarity is the c1fota protein, with a PSSM E-value of 0.0137. Again, this helps to better pigeon-hole the YDR466W protein, allowing the design of more accurate and more effective experiments.

Sanger Institue pfam

The pfam results show a large pkinase domain at the front end of the YDR466W protein. These eukaryotic kinases "are enzymes that belong to a very extensive family of proteins which share a conserved catalytic core common with both serine/threonine and tyrosine protein kinases. There are a number of conserved regions in the catalytic domain of protein kinases. In the N-terminal extremity of the catalytic domain there is a glycine-rich stretch of residues in the vicinity of a lysine residue, which has been shown to be involved in ATP binding. In the central part of the catalytic domain there is a conserved aspartic acid residue which is important for the catalytic activity of the enzyme" (taken from Sanger Institute pkinase data). This again could help to determine which proteins the YDR466W protein interacts with. Also in the pfam results, The pkinase domain is followed by several small regions of low complexity later in the protein. These are insignificant at the moment, but could lead to a further understanding of the protein's structure and function.

SCOP Superfamily

The Superfamily results tried to identify the YDR466W protein with some "superfamilies" of known proteins. This test was extremely productive because of the confidence level that the search produced. It was found that YDR466W belongs to the protein kinase-like superfamily with an E-value of 6.8 e-76. This extremely high level of confidence in the results allows us to identify YDR466W almost definitely as some type of protein kinase, or at least containing that domain.

Protein Structure Initiative

The Protein Structure Initiative is a database that "aims at determination of the 3D structure of all proteins." When searched, it shows that the YDR466W protein has several hits for the presence of the sequence in protein families, but no structural relatives and no other hits for structure information. (See the results here)

Database of Interacting Proteins

This database did not contain any information about the YDR466W protein.

What Is There?

This database lacked information about pathways, enzymes, topics, ortholog clusters, and operon clusters for the YDR466W protein.

Enzymes and Metabolic Pathways Database

This database also lacked information about YDR466W.

Yale Gerstein Lab

The Gerstein Lab database relies on other databases for some of its information, but a great deal of the information comes from research done at Yale. The results for the YDR466W protein show that it has a predicted secondary structure similar to that of the PDB protein 1CKI, a casein kinase I in Rattus norvegicus, and 1APM, a kinase from an unidentified source. The 1CKI and 1APM proteins show many helices and coils, and fewer beta-pleated strands. The 1CKI protein contains two larger beta-pleated regions in addition to two smaller beta regions, while the 1APM protein only has one larger beta-pleated region, but retains the two smaller beta regions. The protein has protein chip data available, which is an exciting development. The protein chip data shows that the YDR466W protein only displayed significant changes in frequency for the phosphorylation of Hsl1 and Axl2 proteins (significant changes = two-fold or greater change in either direction). Also, the substrate specificity test showed that the YDR466W protein showed significant changes in frequency (same criteria as above) for the Hsl1 and Axl2 proteins again. More experiments should be done to determine exactly what the relationship is between these three proteins.

Triples Database

This database contained some information about the YDR466W protein (see the results). It revealed five different clone IDs that could be analyzed for disruption of ORFs and NORFs. These transposon-tagged genes showed different behaviors in the vegetative and sporulation states, and some IDs even contained phenotypic data. However, this phenotypic data showed that the protein displayed wild-type growth patterns in all phenotypically-tested experiments, leading us to believe that YDR466W is not a vital protein.

PDF Protein Interaction Files

These files contained no interactions for the YmL28 protein. MRPL28 (the YmL28 protein gene) and YDR462W (the YmL28 protein ORF) were also searched, but no results were found.

Experiment Design

It would further the knowledge of the YDR466W protein to know the exact crystallized structure of the protein, and also with which proteins YDR466W interacts. The structure of the protein could be analyzed for conserved folds and domains, which would further our knowledge about how sequence affects structure. It could also be analyzed by looking at the 3D structure and determining what proteins it resembles. The proteins that YDR466W interacts with, however, may prove to be even more important. Because the protein shares so much similarity with the kinases, it could be assumed that YDR466W phosphorylates other proteins, meaning that it is upstream of other processes in the cell. This could be analyzed with more protein chip, Y2H, and other proteomic experiments. The results of these experiments could be extremely vital to the understanding of the protein's function.

Links
Peter Lowry's Home Page
Davidson College
Davidson Biology Department
Davidson Genomics Homepage

This page was created by:
Peter Lowry
pelowry@davidson.edu
Davidson College