Mastering Biology Chapter 21
Using BLAST: What Can a Protein Sequence Reveal about Cancer?
[Screenshot from BLAST] Many genes whose normal function is to control cell division can become cancer-causing genes, or oncogenes, through mutation. The normal (nonmutated) versions of these genes are referred to as proto-oncogenes.
In this tutorial, you will investigate a mutation that converts a proto-oncogene into an oncogene. The resulting protein is involved in chronic myelogenous leukemia (CML), a form of cancer that causes the unregulated growth of myeloid cells, bone marrow cells that give rise to white blood cells.
To examine this CML-associated protein, you will use BLAST (Basic Local Alignment Search Tool), a publicly available program for searching known nucleotide and amino acid sequences from several bioinformatics databases organized by the National Center for Biotechnology Information (NCBI). The BLAST algorithm statistically ranks the similarity between an input sequence (called the query) and all other sequences present in a database. The DNA and protein sequences are linked to information about
genes and gene families
the organism the sequence is derived from
the function of the protein (known or predicted)
protein structural information
Thus, BLAST provides a powerful tool for linking gene or protein sequence information with function and biological origin.
Part A - Search for the CML-associated protein sequence in BLAST
The amino acid sequence of the CML-associated protein is shown here:
1 psmafrvhsr ngksytflis sdyeraewre nireqqkkcf rsfsltsvel qmltnscvkl
61 qtvhsiplti nkegeklrvl gynhngewce aqtkngqgwv psnyitpans lekhswyhgp
121 vsrnaaeyll ssgingsflv resesspgqr sislryegrv yhyrintasd gklyvssesr
181 fntlaelvhh hstvadglit tlhypapkrn kptvygvspn ydkwemertd itmkh
This is one of several standard methods used to show the amino acid sequence of a protein; most use the same single-letter abbreviation for each amino acid (e.g., p = proline, s = serine, m = methionine, etc.). Note that in this example, the amino acid sequence is arranged in groups of 10 and that the numbers at the beginning of each line denote the position of the first amino acid in that line in the overall sequence.
This CML-associated protein is known to be the product of a mutation event. Now you will enter this sequence into BLAST and look for similar amino acid sequences in the NCBI databases in order to identify the type of mutation and the genes that are affected.
Go to the BLAST web site by clicking the Launch BLAST button. Then follow the instructions below.
BLAST Search Instructions ( please look attach file for more information )
In the middle of the page under the Basic BLAST heading, click protein blast. A new page will appear.
Copy the amino acid sequence from above (BLAST ignores numbers and spaces) and paste it in the Enter Query Sequence box at the top of the new page.
In the Choose Search Set box, in the pull-down menu to the right of Database, choose the database Non-redundant protein sequence (nr).
In the text box to the right of Organism, type "Homo sapiens," and then click on Homo sapiens (taxid: 9606).
In the Program Selection box, choose blastp (protein-protein BLAST).
In the lower left of the screen (you may need to scroll down), click the BLAST button. Wait for BLAST to complete the search. Initial information (Conserved Domains) may appear quickly, but display of the full results may take 30 seconds. (If your search returns a screen that displays “No significant similarity found,” open Hint 1 to see what might have gone wrong.)
A new screen displays your search results. Briefly scroll down to look at the three main sections, and notice the type of information that is provided in each section. Note that in each of the three sections, similar sequences, or hits, are listed beginning with the best statistical match to your query sequence.
The Graphic Summary gives a color-based summary of the sequence alignment between your query sequence and the most similar sequences (hits). (For this exercise, you can ignore the conserved domains information at the top.)
The Descriptions section provides the unique accession number of each hit, a brief description of the sequence, and several “scores” that quantify the similarity between each hit and your query sequence.
The Alignments section shows the alignment between each hit and your query sequence, amino acid by amino acid.
Scroll to the top of the Graphic Summary section, but ignore the conserved domains information. Place your cursor over the top red bar that represents the first hit. Look at the information in the small text box immediately above the Graphic Summary figure. For an explanation of what appears in the text box, see Hint 2.
[Screenshot from BLAST]
Scroll down to the first entry in the Descriptions section. Notice that the information from the text box above the Graphic Summary matches the information in the Accession, Description, Max score (the same as the S value in the text box), and E value columns.
Scroll back up to the Graphic Summary. Again, place your cursor over the top red bar and click on it. This links you to the information in the Alignments section for this hit. Your hit will appear at the top of the screen beginning with its accession number and description.
Based on the information given in the three different sections, which of the following statement(s) correctly describe(s) the hit that is most similar to the query sequence? Select all that apply.
Based on the information given in the three different sections, which of the following statement(s) correctly describe(s) the hit that is most similar to the query sequence? Select all that apply.
a) It contains a total of 491 amino acids.
b) It is identical to the query sequence in length and amino acid sequence.
c) It is a chimeric protein.
d) Its accession number is CAM33009.1.
Quick Reply
