BIOL368/F14:Chloe Jones Week 8: Difference between revisions
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Chloe Jones (talk | contribs) (→Chapter 6:Working with a single protein sequence: added images) |
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*Program cuts protein at specific site | *Program cuts protein at specific site | ||
<u> Doing primary structure analysis</u> | <u> Doing primary structure analysis</u> | ||
<b> Looking for transmembrane segments</b> | <b> Looking for transmembrane segments</b>[[Image:Transmembrane signals .png|thumb|right|300px|= Looking for transmembrane segments with amino acid scale Hphob. / Kyte & Doolittle.|<b>Figure 4.</b> Looking for transmembrane segments with amino acid scale Hphob. / Kyte & Doolittle.]] | ||
*Used http://web.expasy.org/cgi-bin/protscale/protscale.pl ExPasy Protscale] | *Used http://web.expasy.org/cgi-bin/protscale/protscale.pl ExPasy Protscale] | ||
*Imput accession number: Q75760 | *Imput accession number: Q75760 |
Revision as of 09:49, 21 October 2014
Working with Protein Sequences In-class Activity
Chapter 4: Reading a SWISS-PROT entry
Source: Bioinformatics for Dummies pp.110-123
- Using the database UniProt KB which has two subsections: Swis-Prot and TrEMBL.
- If you search on the keywords "HIV" and "gp120", how many results do you get?
- 180,227 results.
General Information about the entry
- Entry name: 9HIV1
- Primary Accession Number :Q75760
- Secondary Accession Numbers :N/A
- Intergrated into Swiss-Prot on :November 1, 1996
- Sequence was last modified on : November 1, 1996
- Annotations were last modified on : October 1, 2014
Name and origin of the protein
- Protein name:Envelope glycoprotein gp160
- Synonyms:N/A
- Gene name:Env
- From:Homo sapiens (Human) [TaxID: 9606] TaxID:9606
- Taxonomy: Viruses › Retro-transcribing viruses › Retroviridae › Orthoretrovirinae › Lentivirus › Primate lentivirus group
References
- Number of references:9
The comments :?? The cross references :172
- ENA, European Nucleotide Archive (more information possibly)
Keyword
- Apoptosis, fushion of virus membrane with host membrane, host-virus interaction, viral attachment to host cell, viral immunoevasion, Viral penetration into host cytoplasm, virus entry into host cell
Chapter 5: ORFing your DNA sequence
Source: Bioinformatics for Dummies pp.146-147
- Fasta format of Envelope glycoprotein gp160 obtained from UniProt KB was placed in the in the imput box of ORF Finder(Open Reading Frame Finder). 6 parallel horizontal bars were present with integers correlating to the reading frame.
- Putting in DNA sequences: Subject 1 visit 1, clone 1:
- The protein with the open reading frame was in the ORF Finder database was projected to be #2, or #3. Looking at the exPASy database 5’3” Frame 3 had an open reading frame with no stop codons interrupting /truncating the lengh of the DNA.
- Apoptosis, fushion of virus membrane with host membrane, host-virus interaction, viral attachment to host cell, viral immunoevasion, Viral penetration into host cytoplasm, virus entry into host cell
The features Media***
Chapter 6:Working with a single protein sequence
Source: Bioinformatics for Dummies pp.159-195
Predicting the main physico-chemical properties of a protein
- Used the program ExPasy ProtParam for computation of physcial and chemical parameters of a given protein
- Imput the Swiss-Prot/TrEMBL accession number Q75760. Then click compute parameters button >Submit
- Corresponds to the HIV gp120 sequence that was used for the crystal structure for the Huang et al. (2005) paper.
- Can also enter raw sequence
- Save the file
- media ***
Interpreting ProtParam results
- Used ExPasy ProtParam for data.
- Molecular Weight: 96160.4 Daltons
- Extinction Coefficients:
- assuming all pairs of Cys residues form cystines= 184145M1cm1
- assuming all Cys residues are reduced= 182770 M1cm1
- ~Tell you how much light (visible or invisible) your protein absorbs at a certain wavelength.
- Instability
- instability index (II) is computed to be 37.91
- classifies the protein as stable
- Half-Life
- The estimated half-life is:
- 30 hours (mammalian reticulocytes, in vitro)
- >20 hours (yeast, in vivo)
- >10 hours (Escherichia coli, in vivo)
- The estimated half-life is:
Digesting a protein in a computer
- Used ExPasy Peptide Cutter
- Imput accession number: Q75760
- Program cuts protein at specific site
Doing primary structure analysis
Looking for transmembrane segments
- Used http://web.expasy.org/cgi-bin/protscale/protscale.pl ExPasy Protscale]
- Imput accession number: Q75760
- The amino acid scale Hphob. / Kyte & Doolittle was preselected
- Changed the Window size to 19 because best window value for viewing trans membrane regions
- Covert the image to GIF format
- Save file
- Interpreting ProtScale results
- To confirm use Hphob. / Eisenberg et al.scale, set threshold to 1.6
- Piece of paper to over results
- lower paper to strongest peaks visible
- Keep lowering threshold as long as you keep seeing sharp peeks
- 4 Sharp Peaks, 4 Transmembrane domains
- Running TMHMM
- UseTMHMM Q75760 sequence was inputted in FASTA format>Submit
- 5 transmembrane domains identified Figure. #
- Looking for coiled regions
- Used COILS server at EMBnet, input accession number Q75760
- Changed input sequence format to SwissProtID or AC
Predicting post-translational modifications in your protein
- Looking for PROSITE patterns
- Used ScanProsite, input accession number Q75760
- Uncheck excluded motifs with high probability of occurrence, check the excluded profiles from scan box >start the scan
- Interpreting ScanProsite results
- Slide mouse over color rectangles to see name displayed, click to receive more information
- the list of segments contain patterns within protein; numbers indicate position and capital letters are specified and lowercase letters are unspecified by patterns
- ~be careful with short patterns
- Weak signals add up to give strong signals –two related patterns at close distance, significant
- Eliminating weak patterns-multiple sequence alignment
Finding Known Domains in Your Protein
- Finding Domains with InterProScan
- [ http://www.ebi.ac.uk/Tools/pfa/iprscan5/ InterProScan], imput fasta format of Q75760 sequence>submit
- [ http://www.ebi.ac.uk/Tools/pfa/iprscan5/ InterProScan], imput fasta format of Q75760 sequence>submit