00:01
So what we know is that we have a peptide that is being acid hydrolyzed and the hydrolyzed sample is then applied to ion exchange chromatography at doex50.
00:17
And what we know about this is that doex50 is a cation exchange chromatography, which means it has a carry negative charge and it binds to positively charged molecules.
00:48
Now let's take a look at the peptide.
00:51
So the peptide has been digested and then obviously it has six amino acid and it will be broken down into five different amino acid because you have two same amino acids which is a histidine.
01:05
So you have glutamine and you have histidine, you have leucine, valine, and then you have lysine.
01:24
So it's very important for us to find out the pi of each amino acid.
01:30
So let's see the pi of glutamate acid is in fact 3 .2.
01:42
Then histidine has a pi of 7 .59.
01:52
Then leucine has a pi of 5 .98.
02:01
And then valine has a pi of 5 .96.
02:12
And lastly lysine has a pi of 9 .74.
02:21
Now doex50 cation exchange chromatography is done at a ph about 7.
02:27
So we want to find out the charge each of amino acid carry when the ph is 7.
02:35
So what we know is that when an amino acid is sitting in a ph that is higher than its pi, it carries a negative charge.
02:50
Let me write that.
03:20
So molecule carries negative charge when sitting at a ph above the pi and on the other hand a positive charge when sitting at a ph below its pi.
03:38
So you can see that for glu, glutamate acid has a pi of 3 .2.
03:44
So it's sitting at ph 7 which is above the pi.
03:48
So it carry negative charge.
03:52
And histidine 7 .59 is higher than 7.
03:56
So it's sitting at a ph below its pi.
03:59
Positive charge.
04:01
But because it's only slightly higher than the 7, so it carry only a few positive charge.
04:14
And next one, leucine 5 .98 sits at ph 7 which is above its pi.
04:23
So negative charge.
04:25
And because it's closer to 7, so it actually carry less negative charge than glutamic acid...