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Organic : Amino Acid Analysis
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 Message 1 of 3 in Discussion 
From: MSN NicknameGuy_SoCa  (Original Message)Sent: 10/21/2008 9:06 PM
Hi Steve and all!
I am trying to work this problem out and I'm only partially putting all the information together but I'm still missing how to integrate part (d) and how to chose from the amino acids that don't contribute to the net charge of the oligopeptide.
Thanx in advanced!

You have isolated a new pentapeptide from a bacterium. A series of experiments reveals the following about your peptide:

a) The column elution profile from amino acid analysis of the pentapeptide yields five distinct peaks;
b) the pentapeptide strongly absorbs light at 280nm
c) the pentapeptide has a net charge of " -0.5 " at pH 10.5
d) bubbling oxygen into the petapeptide solution for an extended period of time result in decapenptides being formed.

What are the amino acids making up this oligopeptide?


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Reply
 Message 2 of 3 in Discussion 
From: MSN Nickname·Steve·Sent: 10/21/2008 10:40 PM
The information limits the choices of amino acids, especially the �?.5 overall charge.  From the given information, it can be concluded that,
 
a)  Five different amino acids are present.
b)  One or more aromatic R-groups are present (Trp, Tyr, or Phe).
c)  Charges of all R-groups plus the C-terminus add up to �?.5.
d)  Oxidation of cysteine R–SH –�?gt; cystine R–S–S–R (the decapeptide)  +  2 H+  +  2e�?/SUP>
     (so cysteine is present).

If the pH is significantly above pKa, then the group will be in its deprotonated form,
(–COOH –�?gt; –COO�?/SUP>  and  –NH3+ –�?gt; –NH2) with corresponding charges.
 
Amino Acids with Non-Neutral R-Groups:
Amino Acid       Approximate pKa of R-group       Charge of R-group at pH 10.5                   
Asp                                   3.9                                         �?
Glu                                    4.1                                         �?
Arg                                  12.5                                         +1
Lys                                  10.5                                           50% +1, 50% 0  (pH = pKa)
His                                    6.0                                           0
Cys                                   8.4 (–SH)                                �?
Tyr                                   10.5                                           50% 0, 50% �? (pH = pKa)
 

N-terminus    pKa = 8.0, charge at pH 10.5 = 0
C-terminus    pKa = 3.0, charge at pH 10.5 = �?
 

At pH 10.5, the following adds up to �?.5:
(C-terminus)     �?
Cys                 �?
Arg                  +1
Lys                  +0.5  (50% +1, 50% 0)
Total Charge     �?.5

Cysteine is definitely present.  The cysteine R-group at pH 10.5 has a �? charge (R–S�?/SUP>).  The C-terminus will also be in its COO�?/SUP> form at pH 10.5, so that's another �? charge, bringing the total to �? at this point.  To get the charge to �?.5, there is not any choice but to have an arginine with the +1 guanadino group at pH 10.5, which brings the overall charge down to �?, plus a lysine which is 50% protonated (–NH3+) at pH 10.5, netting to a +0.5 charge, which makes the overall charge �?.5.

To bring the total number of amino acids up to five, we need two more with neutral R-groups.  One at least of these will contain an aromatic ring, such as phenylalanine or tryptophan, both which absorbs light at 280 nm.  Since it states that the peptide strongly absorbs at 280 nm, perhaps both of these are present.  Otherwise, I can't tell how to determine what the fifth amino acid is from the given information. 
 

Steve
 
 

Amino Acids Structures and pKa Values:
http://www.cem.msu.edu/~cem252/sp97/ch24/ch24aa.html

Reply
 Message 3 of 3 in Discussion 
From: MSN NicknameGuy_SoCaSent: 10/21/2008 11:35 PM
Sweet!
Thanks Steve