To predict signal peptide in eukaryotic sequence

EXPERIMENT#10

OBJECT:-

To predict signal peptide in eukaryotic sequence.

Theory:-

A signal peptide (some time referred to as signal sequence, targeting signal, localization signal ,transit peptide , leader sequence or leader peptide)is a short (5-30 amino acid)peptide present at the N terminal of the majority of newly synthesized proteins that are destined towards the secretary pathway. These proteins include those that reside either inside certain organelles (endoplasmic reticulum golgi bodies or   endosomes),secreted from the cells or inserted into most cellular membranes. In eukaryotes , signal sequences direct the insertion of proteins into the membrane of the endoplasmic reticulum and are usually cleaved off by signal peptidase. The resulting signal peptide are presumably rapidly degraded but some still have function on their own. SignalP 4.1 server predicts the presence and location of signal peptide cleavage sites in amino acid sequences from different organisms: Gram-positive prokaryotes, Gram-negative prokaryotes, and eukaryotes. The method incorporates a prediction of cleavage sites and a signal peptide/non-signal peptide prediction based on a combination of several artificial neural networks. Organism group:
It is important for performance that you choose the correct organism group — Eukaryotes, Gram-negative bacteria or Gram-positive bacteria — since the signal peptides of these three groups are known to differ from each other.
Gram-positive bacteria correspond to Actinobacteria and Firmicutes in the NCBI Taxonomy.
Gram-negative bacteria are all other eubacteria, except Tenericutes (including Mycoplasma), which seem to lack a type I signal peptidase and therefore do not have standard signal peptides You can see which cutoff values are being used in the boxes marked "D-cutoff". They will change if you change the setting for "D-cutoff values" or "Organism group".
If you want to experiment with your own cutoff values, select "User defined" and the boxes will go blank, ready for you to fill in values between 0 and 1.

PROCEDURE:-

1.In Expasy signal peptide was selected.

2. Fasta sequence of protein was pasted in input box.

3.After tat group of organism was selected (gram negative, gram positive, eukaryotes)

4.Result was noted.

Dicussion:-

In this experiment we predicted signal peptide in eukaryotic sequence, by using expasy  search tool, for that purpose , we pasted protein sequence of a eukaryotic organism in a bar of search and found a graph, we observed a peak (blue colored), it indicated site of proteolytic cleavage. In graph green line showed hydrphobicity residues.

We have to consider those lines which are above the base-line. Green and blue line played basic role in signal peptide.

We observed cleavage site between 23 and 24^th amino acids ,it indicates that signal peptide is present in given sequence. Signal peptide helps the protein to reach its destination, than through cleavage it cleaved out from protein.

Discussion:-

In this experiment we predicted signal peptide in eukaryotic sequence, by using expasy  search tool, for that purpose , we pasted protein sequence of a eukaryotic organism in a bar of search and found a graph, we observed a peak (blue colored), it indicated site of proteolytic cleavage. In graph green line showed hydrphobicity residues.

Any thing above cut off line is significant. The software looked for 3 parameters(1.short stretch of hydrophobic amino acids,2.short peptide,3.cleavage site).It only look 70 begining aminoacids because if sinal peptide present it would be in begining.In translated protein Met is first amino acid but in functional it is not the first aminoacid. By looking the graph we can predict the presence and absence of signal peptide.If signal peptide not present reason might be that protein may be cytoplasmic or there is only functional sequence uploaded or may be there is partial sequence not complete sequence.