Protein Analysis
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Characterize the sequence of the plant enzyme Alcohol dehydrogenase.

  Sample User Question
Analysis/Comments
Flow Chart
Step By Step Guide
Additional Notes 		 

Sample User Question back to top

Characterize the sequence of the plant enzyme Alcohol dehydrogenase.
  • What plant species has the enzyme Alcohol dehydrogenase been isolated from?
  • Apart from the Zinc-containing alcohol dehydrogenases signatures (PROSITE link), how many and what other protein motifs are found in the sequence?
  • What amino acids does the Zinc atom bind to?
User Questions: Proteomic tools
  • Can the protein be cut by Trypsin?
  • Which patterns are more specific for this protein?
  • What are the resulting peptid masses from a tryptic digestion (approximatelly)?
  • What is the Hydrophobicity profile according to Kyte & Doolittle?
  • What is the HPLC retention at a physiological pH?

Analysis/Comments back to top

Flow Chart back to top

  1. ExPASY - The ExPASy (Expert Protein Analysis System) proteomics server of the Swiss Institute of Bioinformatics (SIB).

  2. Sequence Retrieval System (SRS) - A Network Browser for Databanks in Molecular Biology.

  3. ScanProsite - Scan a protein sequence for the occurrence of patterns and profiles stored in the PROSITE database.

  4. PeptideCutter - Predicts potential protease and cleavage sites and sites cleaved by chemicals in a given protein sequence.

  5. PeptideMass - - Calculate masses of peptides and their post-translational modifications for a SWISS-PROT or TrEMBL entry or for a user sequence.

  6. ProtScale - Amino acid scale representation (Hydrophobicity, other conformational parameters, etc.).

Step By Step Guide back to top

ExPASY

  • Access the Sequence Retrieval System (SRS) via one of the databases home pages (e.g., SWISS-PROT and TrEMBL)
    • Start a new session (click start).
    • Choose the databases SWISS-PROT and TrEMBL, click continue.
    • Search for alcohol[All Text], dehydrogenase[All Text] and plant[Organism], click Do Query.

Result:
  • 1 Entry found: Click SWISS_PROT:MTD_MESCR to view NiceProt View of SWISS-PROT: P93257
  • Plant: Mesembryanthemum crystallinum (Common ice plant)
  • Features field: Clicking on the numbers in the from...to column will bring up the aa sequence with the aa high-lighted in red (Cs and H).
ScanProsite

  • Scroll down the Entry P93257 to ScanProsite and click it.
  • The program is already set up for the search (Acc.no is provided when coming from an entry page). Click Start the Scan at the left bottom of the page.
Result:
  • Seven different patterns could be found in the sequence.
PeptideCutter

  • Scroll down the Entry P93257 to Tools: PeptideCutter and click it.
  • The program is already set up for the search (Acc.no is provided when coming from an entry page). Check the radio button for "only the following selection of enzymes and chemicals" and check the box for Trypsin.
Result:
  • 33 different Trypsin cleavage sites are found in the sequence.
PeptideMass

  • Scroll down the Entry P93257 to Tools: PeptideMass and click it.
  • The program is already set up for the search (Acc.no is provided when coming from an entry page). Leave the default parameters checked (enzyme: Trypsin) and click Perform the cleavage.
Result:
  • The next page shows the masses of the cleavage products in a table.
ProtScale

  • Scroll down the Entry P93257 to Tools: ProtScale and click it.
  • The program is already set up for the search (Acc.no is provided when coming from an entry page). Leave the default parameters checked (Hphob. / Kyte & Doolittle) and click Submit.
Result:
  • The next page shows the Hydrophobicity profile according to Kyte & Doolittle.
  • Use the same program to retrieve the HPLC retention at pH 7.4.

Additional Notes back to top

  • 3D information can be accessed via cross-reference to Pfam (Protein families database of alignments and HMMs).



Protein/User Question 1 Return to Question Slide Revised 07/30/2003