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Isoelectric point of aspartate

Isoelectric point of aspartate


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On the 3rd diagram (from left), the one with net charge = -1 ,

Why did it lose the H to OH- from the CH2CO2H from the neutral form ?

I was told that while moving to the right , the OH- will take H+ electrons meaning it should take H+ from NH3… ?

Is there any reason why they took the H from CH2CO2H ?

Any help would be appreciated!


The -CO₂H groups are carboxylic acid groups, better represented as -COOH. These, being acidic, have a lower pKₐ than the -NH₃⁺ group and so as the pH increases these -COOH groups lose their protons first.

The pKₐ values for the three are shown below the arrows: as you can see the α carboxylic acid is more acidic than that in the side chain, but they are both much more acidic than the protonated amino group.


Titration of Aspartate with Hydroxide

On the left in both the chemical reaction and the titration curve, you should imagine that aspartic acid is in a very acidic solution at a pH of about 0. As you move from left to right accross the page, you are adding hydroxide to the solution. This increases the hydroxide concentration and decreases the hydrogen ion concentration. Note that aspartate loses protons as you move from left to right. At the first pKa, the alpha-carboxyl dissociates. At the second pKa, the R-group carboxyl dissociates, At the third pKa, the alpha-amino dissociates.

Note that at each pKa, the solution is buffered. That is, it resists changes in pH as hydroxide is added. Also note, that the pI occurs where aspartate has no net charge.

  • amino acids without any charged R-group (alanine, glycine, . )
  • lysine and arginine
  • aspartate and glutamate
  • histidine

The isoelectric point is the pH at which an amino acid or protein has no net charge and will not migrate towards the anode or cathode in an electric field. The charges on any amino acid at a given pH are a function of their pKas for dissociation of a proton from the alpha-carboxyl groups, the alpha-amino groups, and the side chains (R-group). The pKa for the alpha-amino groups and the alpha-carboxyl groups are about 2 and 10 (Figure 6.1). The pKas of the important side chains are shown in Figure 6.9.

You start by having a very rough idea of the structure of the amino acid. What are the acidic groups and what are their pKas. Next, you try to visualize the amino acid fully associated with hydrogen and what the charge on the molecule would be. Next, you visualize removing hydrogen ions by titrating with hydroxide ions. You will remove hydrogen ions from the group with the lowest pKa first and, then from the next higher pKa. Eventually you reach the pI.

At very acidic pHs, the R-group is in the COOH form, the alpha-amino group is in the –NH3 + form and the alpha-carboxyl group is in the COOH form so aspartate has a net charge of +1

As we titrate with hydroxide ion, we remove hydrogen ions. They combine with hydroxide ions and become water. When we reach pH 2, the protons on half the alpha-carboxyl groups are removed. This is not the pI because half of the alpha-carboxyl have a negative charge but all of the alpha-amino groups –NH3 + have a positive charge and all of the R-Group (COOH ) have no charge. So, the net charge on the aspartate molecules is a positive 0.5.

As we titrate with more hydroxide ions, we reach a point half way between pKa1 and pKa2. At this pH, half of the protons have been removed from the two carboxyl groups and half of the carboxyl groups are not dissociated so the net charge on the carboxyl groups is a -1. The alpha-amino group is fully charged so it has a net charge of +1. The net charge on the aspartate molecules is 0. This is the pI

As we titrate with more hydroxide ions, we reach the pKa2, At this pH, all of the protons have been removed from the alpha-carboxyl group and half of the half of the protons have been removed from the R-group carboxyl groups. The net charge on the carboxyl groups is a -1.5. The alpha-amino group is fully charged so it has a net charge of +1. The net charge on the aspartate molecules is -0.5.

As we titrate with more hydroxide ions, we reach the pKa3, a point where all the carboxyl groups are dissociated and only half of the alpha-amino groups still have a positive charge. The net charge is a negative 1.5. We did not have to go this far to determine the pI but I thought it might be useful.


A Corner of Structural Biology

Let’s start from isoelectric point definition:

Isoelectric point (pI) is a pH in which net charge of protein is zero. In case of proteins isoelectric point mostly depends on seven charged amino acids: glutamate (δ-carboxyl group), aspartate (ß-carboxyl group), cysteine (thiol group), tyrosine (phenol group), histidine (imidazole side chains), lysine ( ε -ammonium group) and arginine (guanidinium group). Additonally, one should take into account charge of protein terminal groups (NH2 i COOH). Each of them has its unique acid dissociation constant referred to as pK.
Moreover, net charge of the protein is in tight relation with the solution (buffer) pH. Keeping in main this we can use Henderson-Hasselbach equationto calculate protein charge in certain pH:

– for negative charged macromolecules:

– for positive charged macromolecules:

* Arg was not included in the study and the average pK from all other scales was taken

More advanced algorithm, implemented in ProMoST, takes into account localization of the charched amino acid:

aa N term middle C term
K
R
H
D
E
C
U*
Y
10.00
11.50
4.89
3.57
4.15
8.00
5.20
9.34
9.80
12.50
6.08
4.07
4.45
8.28
5.43
9.84
10.30
11.50
6.89
4.57
4.75
9.00
5.60
10.34
* pK was taken from Byun et al. 2011

Additionally different pK values are used for N and C terminus depending on uncharched amino acid if aplicable:

aa N C aa N C aa N C aa N C
G
A
S
P
7.50
7.58
6.86
8.36
3.70
3.75
3.61
3.61
V
T
I
L
7.44
7.02
7.48
7.46
3.69
3.57
3.72
3.73
N
Q
M
F
7.22
6.73
6.98
6.96
3.64
3.57
3.68
3.98
W
X*
Z**
B***
7.11
7.26
6.96
7.46
3.78
3.57
3.54
3.57
* X – average from all amino acids
** Z =(E+Q)/2
*** B =(N+D)/2

Now, having this few peaces of information we can try to write simple computer program which calculate isoelectric point. We will use free compiler DevC++ as the program will be written in C++ programming language. To read next section you should have at least basic knowledge in C++.


Amino Acid

To form a molecule with its functional groups, having a positive and negative charge.

1 ml Ninhydrin on 1 ml protein solution shows violet color after heating. It shows the presence of alpha-amino acids.

Sanger reagent reacts with an amino group in a mild alkaline medium under cold conditions.

Reacts with the Amino group to release nitrogen and form the corresponding hydroxyl.

Classification based on polarity

  1. Polar Amino Acids
  2. Non-polar Amino Acids
  • Polar amino acids:
  1. In this category there are 11 amino acids listed down:
  2. Polar Uncharged: Serine, Threonine, Cysteine, Asparagine, Glutamine, and Tyrosine.
  3. Polar Charged: Histidine, Lysine, Arginine, Aspartate, and Glutamate
  • Non-polar amino acid
  1. In this category there are 9 amino acids:
  2. Glycine, Alanine, Proline, Valine, Leucine, Isoleucine, Tryptophan, Phenylalanine, and Methionine.

Essential and non-Essential amino acids

based on the requirement of our body:

  • What are the Essential amino acids?
  1. Out of 20 amino acids, there are 9 are in the list of essential amino acids. We need to take these amino acids from outside (food sources).
  2. Isoleucine, Valine, Lysine, Phenylalanine, Methionine, Threonine and Tryptophan
  • What are the Non-essential amino acids?
  1. These amino acids can be made by our body.
  2. Arginine, Cysteine, Glutamine, Tyrosine, Glycine, Proline, Serine, Alanine, Aspartate, and Asparagine.

What is the isoelectric point?

The pH when the total charge of an amino acid is zero, known as an isoelectric point.


Ceramic materials

The isoelectric points (IEP) of metal oxide ceramics are used extensively in material science in various aqueous processing steps (synthesis, modification, etc.). In the absence of chemisorbed or physisorbed species particle surfaces in aqueous suspension are generally assumed to be covered with surface hydroxyl species, M-OH (where M is a metal such as Al, Si, etc.). ⎚] At pH values above the IEP, the predominate surface species is M-O − , while at pH values below the IEP, M-OH2 + species predominate. Some approximate values of common ceramics are listed below: ⎛] ⎜]

Material IEP Material IEP Material IEP Material IEP Material IEP Material IEP
WO3 ⎝] 0.2-0.5 Ta2O5 ⎝] 2.7-3.0 δ-MnO2 1.5 Fe2O3 ⎝] 3.3-6.7 Fe2O3 ⎝] 8.4-8.5 ZnO ⎝] 8.7-10.3
Sb2O5 ⎝] <0.4-1.9 SnO2 ⎞] 4-5.5 (7.3) β-MnO2 ⎟] 7.3 CeO2 ⎝] 6.7-8.6 α Al2O3 8-9 NiO ⎞] 10-11
V2O5 ⎝] ⎟] 1-2 (3) ZrO2 ⎝] 4-11 TiO2 ⎠] 2.8-3.8 Cr2O3 ⎝] ⎟] 6.2-8.1 (7) Si3N4 ⎞] 9 PbO ⎝] 10.7-11.6
SiO2 ⎝] 1.7-3.5 MnO2 4-5 Si3N4 6-7 γ Al2O3 7-8 Y2O3 ⎝] 7.15-8.95 La2O3 10
SiC ⎡] 2-3.5 ITO ⎢] 6 Fe3O4 ⎝] 6.5-6.8 Tl2O ⎣] 8 CuO ⎞] 9.5 MgO ⎝] 12-13 (9.8-12.7)

Note: The following list gives the isoelectric point at 25 °C for selected materials in water. The exact value can vary widely, depending on material factors such as purity and phase as well as physical parameters such as temperature. Moreover, the precise measurement of isoelectric points can be difficult, thus many sources often cite differing values for isoelectric points of these materials.

Mixed oxides may exhibit isoelectric point values that are intermediate to those of the corresponding pure oxides. For example, a synthetically prepared amorphous aluminosilicate (Al2O3-SiO2) was initially measured as having IEP of 4.5 (the electrokinetic behavior of the surface was dominated by surface Si-OH species, thus explaining the relatively low IEP value). ⎤] Significantly higher IEP values (pH 6 to 8) have been reported for 3Al2O3-2SiO2 by others. ⎞] Similarly, also IEP of barium titanate, BaTiO3 was reported in the range 5-6 ⎞] while others got a value of 3. ⎥] Mixtures of titania (TiO2) and zirconia (ZrO2) were studied and found to have an isoelectric point between 5.3-6.9, varying non-linearly with %(ZrO2). ⎦] The surface charge of the mixed oxides was correlated with acidity. Greater titania content led to increased Lewis acidity, whereas zirconia-rich oxides displayed Br::onsted acidity. The different types of acidities produced differences in ion adsorption rates and capacities.


What is isoelectric pH of amino acid?

The word isoelectric or isoelectronic comes from 'iso,' which means the same, and 'electric,' which implies charge. The isoelectric point or pI of an amino acid is the pH at which an amino acid has a net charge of zero.

Furthermore, what is pI in pH? The isoelectric point (pI, pH(I), IEP), is the pH at which a molecule carries no net electrical charge or is electrically neutral in the statistical mean. The standard nomenclature to represent the isoelectric point is pH(I), although pI is also commonly seen, and is used in this article for brevity.

Keeping this in consideration, how does isoelectric point relate to pH?

4.6 Isoelectric Point Precipitation The isoelectric point (pI) is the pH of a solution at which the net charge of a protein becomes zero. At solution pH that is above the pI, the surface of the protein is predominantly negatively charged, and therefore like-charged molecules will exhibit repulsive forces.

Key Takeaways: pKa Definition The pKa value is one method used to indicate the strength of an acid. pKa is the negative log of the acid dissociation constant or Ka value. A lower pKa value indicates a stronger acid. That is, the lower value indicates the acid more fully dissociates in water.


RESULTS

Database use

The Proteome-pI database incorporates multiple browsing and searching tools. First, it can be searched and browsed by organism name, average isoelectric point, molecular weight or amino acid frequencies (see also Table 2). Proteins with extreme pI values are also available. For individual proteomes, users can retrieve proteins of interest given the method, isoelectric point and molecular weight ranges (this particular feature can be highly useful to limit potential targets in analysis of 2D-PAGE gels or before conducting mass spectrometry). Additionally, precalculated fractions of proteins according to isoelectric point are also available. Finally, some general statistics (total number of proteins, amino acids, average sequence length, amino acid frequency) and links to other databases (UniProt, NCBI) can be found (see Figure 1 for an example).

Proteome-pI example report for Salmonella enterica. At the top, the average isoelectric point, precalculated fractions of proteins according to isoelectric point and virtual 2D-PAGE plot for the proteome are shown. In the next section, the user can retrieve a subset of proteins within specified isoelectric point and molecular weight ranges calculated using a particular method. Next, proteins with minimal and maximal isoelectric points are presented along with some general statistics.

Proteome-pI example report for Salmonella enterica. At the top, the average isoelectric point, precalculated fractions of proteins according to isoelectric point and virtual 2D-PAGE plot for the proteome are shown. In the next section, the user can retrieve a subset of proteins within specified isoelectric point and molecular weight ranges calculated using a particular method. Next, proteins with minimal and maximal isoelectric points are presented along with some general statistics.

Amino acid frequency for the kingdoms of life in the Proteome-pI database

Kingdom . Ala . Cys . Asp . Glu . Phe . Gly . His . Ile . Lys . Leu . Met . Asn . Pro . Gln . Arg . Ser . Thr . Val . Trp . Tyr . Total amino acids .
Viruses 6.61 1.76 5.81 6.04 4.25 5.79 2.15 6.53 6.35 8.84 2.46 5.41 4.62 3.39 5.24 7.06 6.06 6.50 1.19 3.94 6 150 189
Archaea 8.20 0.98 6.21 7.69 3.86 7.58 1.77 7.03 5.27 9.31 2.35 3.68 4.26 2.38 5.51 6.17 5.44 7.80 1.03 3.45 89 488 664
Bacteria 10.06 0.94 5.59 6.15 3.89 7.76 2.06 5.89 4.68 10.09 2.38 3.58 4.61 3.58 5.88 5.85 5.52 7.27 1.27 2.94 3 716 982 916
Eukaryota 7.63 1.76 5.40 6.42 3.87 6.33 2.44 5.10 5.64 9.29 2.25 4.28 5.41 4.21 5.71 8.34 5.56 6.20 1.24 2.87 3 743 221 293
All 8.76 1.38 5.49 6.32 3.87 7.03 2.26 5.49 5.19 9.68 2.32 3.93 5.02 3.90 5.78 7.14 5.53 6.73 1.25 2.91 7 555 843 062
Kingdom . Ala . Cys . Asp . Glu . Phe . Gly . His . Ile . Lys . Leu . Met . Asn . Pro . Gln . Arg . Ser . Thr . Val . Trp . Tyr . Total amino acids .
Viruses 6.61 1.76 5.81 6.04 4.25 5.79 2.15 6.53 6.35 8.84 2.46 5.41 4.62 3.39 5.24 7.06 6.06 6.50 1.19 3.94 6 150 189
Archaea 8.20 0.98 6.21 7.69 3.86 7.58 1.77 7.03 5.27 9.31 2.35 3.68 4.26 2.38 5.51 6.17 5.44 7.80 1.03 3.45 89 488 664
Bacteria 10.06 0.94 5.59 6.15 3.89 7.76 2.06 5.89 4.68 10.09 2.38 3.58 4.61 3.58 5.88 5.85 5.52 7.27 1.27 2.94 3 716 982 916
Eukaryota 7.63 1.76 5.40 6.42 3.87 6.33 2.44 5.10 5.64 9.29 2.25 4.28 5.41 4.21 5.71 8.34 5.56 6.20 1.24 2.87 3 743 221 293
All 8.76 1.38 5.49 6.32 3.87 7.03 2.26 5.49 5.19 9.68 2.32 3.93 5.02 3.90 5.78 7.14 5.53 6.73 1.25 2.91 7 555 843 062

*Similar statistics for all 5029 proteomes included in Proteome-pI are available online on individual subpages. For di-amino acid frequencies see Supplementary Table S2 .

Kingdom . Ala . Cys . Asp . Glu . Phe . Gly . His . Ile . Lys . Leu . Met . Asn . Pro . Gln . Arg . Ser . Thr . Val . Trp . Tyr . Total amino acids .
Viruses 6.61 1.76 5.81 6.04 4.25 5.79 2.15 6.53 6.35 8.84 2.46 5.41 4.62 3.39 5.24 7.06 6.06 6.50 1.19 3.94 6 150 189
Archaea 8.20 0.98 6.21 7.69 3.86 7.58 1.77 7.03 5.27 9.31 2.35 3.68 4.26 2.38 5.51 6.17 5.44 7.80 1.03 3.45 89 488 664
Bacteria 10.06 0.94 5.59 6.15 3.89 7.76 2.06 5.89 4.68 10.09 2.38 3.58 4.61 3.58 5.88 5.85 5.52 7.27 1.27 2.94 3 716 982 916
Eukaryota 7.63 1.76 5.40 6.42 3.87 6.33 2.44 5.10 5.64 9.29 2.25 4.28 5.41 4.21 5.71 8.34 5.56 6.20 1.24 2.87 3 743 221 293
All 8.76 1.38 5.49 6.32 3.87 7.03 2.26 5.49 5.19 9.68 2.32 3.93 5.02 3.90 5.78 7.14 5.53 6.73 1.25 2.91 7 555 843 062
Kingdom . Ala . Cys . Asp . Glu . Phe . Gly . His . Ile . Lys . Leu . Met . Asn . Pro . Gln . Arg . Ser . Thr . Val . Trp . Tyr . Total amino acids .
Viruses 6.61 1.76 5.81 6.04 4.25 5.79 2.15 6.53 6.35 8.84 2.46 5.41 4.62 3.39 5.24 7.06 6.06 6.50 1.19 3.94 6 150 189
Archaea 8.20 0.98 6.21 7.69 3.86 7.58 1.77 7.03 5.27 9.31 2.35 3.68 4.26 2.38 5.51 6.17 5.44 7.80 1.03 3.45 89 488 664
Bacteria 10.06 0.94 5.59 6.15 3.89 7.76 2.06 5.89 4.68 10.09 2.38 3.58 4.61 3.58 5.88 5.85 5.52 7.27 1.27 2.94 3 716 982 916
Eukaryota 7.63 1.76 5.40 6.42 3.87 6.33 2.44 5.10 5.64 9.29 2.25 4.28 5.41 4.21 5.71 8.34 5.56 6.20 1.24 2.87 3 743 221 293
All 8.76 1.38 5.49 6.32 3.87 7.03 2.26 5.49 5.19 9.68 2.32 3.93 5.02 3.90 5.78 7.14 5.53 6.73 1.25 2.91 7 555 843 062

*Similar statistics for all 5029 proteomes included in Proteome-pI are available online on individual subpages. For di-amino acid frequencies see Supplementary Table S2 .

Moreover, apart from the data for individual proteomes, one can also obtain precalculated isoelectric points from all major protein databases, including nr ( 21), UniProt, PDB ( 22) and SwissProt ( 23) (more details in Supplementary Data).


Isoelectric point of aspartate - Biology

Amino Acids

Proteins are formed by polymerizing monomers that are known as amino acids because they contain an amine (-NH2) and a carboxylic acid (-CO2H) functional group. With the exception of the amino acid proline, which is a secondary amine, the amino acids used to synthesize proteins are primary amines with the following generic formula.

These compounds are known as a -amino acids because the -NH2 group is on the carbon atom next to the -CO2H group, the so-called carbon atom of the carboxylic acid.

The chemistry of amino acids is complicated by the fact that the -NH2 group is a base and the -CO2H group is an acid. In aqueous solution, an H + ion is therefore transferred from one end of the molecule to the other to form a zwitterion (from the German meaning mongrel ion, or hybrid ion).

Zwitterions are simultaneously electrically charged and electrically neutral. They contain positive and negative charges, but the net charge on the molecule is zero.

More than 300 amino acids are listed in the Practical Handbook of Biochemistry and Molecular Biology, but only the twenty amino acids in the table below are used to synthesize proteins. Most of these amino acids differ only in the nature of the R substituent. The standard amino acids are therefore classified on the basis of these R groups. Amino acids with nonpolar substituents are said to be hydrophobic (water-hating). Amino acids with polar R groups that form hydrogen bonds to water are classified as hydrophilic (water-loving). The remaining amino acids have substituents that carry either negative or positive charges in aqueous solution at neutral pH and are therefore strongly hydrophilic.

The 20 Standard Amino Acids

NAME STRUCTURE
(AT NEUTRAL pH)
Nonpolar (Hydrophobic) R Groups
Glycine (Gly)
Alanine (Ala)
Valine (Val)
Leucine (Leu)
Isoleucine (Ile)
Proline (Pro)
Methionine (Met)
Phenylalanine (Phe)
Tryptophan (Trp)
Polar (Hydrophilic) R Groups
Serine
(ser)
Threonine
(Thr)
Tyrosine
(Tyr)
Cysteine
(Cys)
Asparagine
(Asn)
Glutamine
(Gln)
Negatively Charged R Groups
Aspartic acid (Asp)
Glutamic acid
(Glu)
Positively Charged R Groups
Lysine
(Lys)
Arginine
(Arg)
Histidine
(His)

Use the structures of the following amino acids in the table of standard amino acids to classify these compounds as either nonpolar/hydrophobic, polar/hydrophilic, negatively charged/hydrophilic, or positively charged/hydrophilic.

Amino Acids as Stereoisomers

With the exception of glycine, the common amino acids all contain at least one chiral carbon atom. These amino acids therefore exist as pairs of stereoisomers. The structures of the D and L isomers of alanine are shown in the figure below. Although D amino acids can be found in nature, only the L isomers are used to form proteins. The D isomers are most often found attached to the cell walls of bacteria and in antibiotics that attack bacteria. The presence of these D isomers protects the bacteria from enzymes the host organism uses to protect itself from bacterial infection by hydrolyzing the proteins in the bacterial cell wall.

A few biologically important derivatives of the standard amino acids are shown in the figure below. Anyone who has used an "anti-histamine" to alleviate the symptoms of exposure to an allergen can appreciate the role that histamine a decarboxylated derivative of histidine plays in mediating the body's response to allergic reactions. L-DOPA, which is a derivative of tyrosine, has been used to treat Parkinson's disease. This compound received notoriety a few years ago in the film Awakening, which documented it's use as a treatment for other neurological disorders. Thyroxine, which is an iodinated ether of tyrosine, is a hormone that acts on the thyroid gland to stimulate the rate of metabolism.

Acetic acid and ammonia often play an important role in the discussion of the chemistry of acids and bases. One of these compounds is a weak acid the other is a weak base.

Thus, it is not surprising that an H + ion is transferred from one end of the molecule to the other when an amino acid dissolves in water.

The zwitterion is the dominant species in aqueous solutions at physiological pH (pH 7). The zwitterion can undergo acid-base reactions, howeer, if we add either a strong acid or a strong base to the solution.

Imagine what would happen if we add a strong acid to a neutral solution of an amino acid in water. In the presence of a strong acid, the -CO2 - end of this molecule picks up an H + ion to form a molecule with a net positive charge.

In the presence of a strong base, the -NH3 + end of the molecule loses an H + ion to form a molecule with a net negative charge.

The figure below shows what happens to the pH of an acidic solution of glycine when this amino acid is titrated with a strong base, such as NaOH.

In order to understand this titration curve, let's start with the equation that describes the acid-dissociation equilibrium constant expression for an acid, HA.

Let's now rearrange the Ka expression,

take the log to the base 10 of both sides of this equation,

and then multiply both sides of the equation by -1.

By definition, the term on the left side of this equation is the pH of the solution and the first term on the right side is the pKa of the acid.

The negative sign on this right side of this equation is often viewed as "inconvenient." The derivation therefore continues by taking advantage of the following feature of logarithmic mathematics

to give the following form of this equation.

This equation is known as the Henderson-Hasselbach equation, and it can be used to calculate the pH of the solution at any point in the titration curve.

The following occurs as we go from left to right across this titration curve.

  • The pH initially increases as we add base to the solution because the base deprotonates some of the positively charged H3N + CH2CO2H ions that were present in the strongly acidic solution.
  • The pH then levels off because we form a buffer solution in which we have reasonable concentrations of both an acid, H3N + CH2CO2H, and its conjugate base, H3N + CH2CO2 - .
  • When virtually all of the H3N + CH2CO2H molecules have been deprotonated, we no longer have a buffer solution and the pH rises rapidly when more NaOH is added to the solution.
  • The pH then levels off as some of the neutral H3N + CH2CO2 - molecules lose protons to form negatively charged H2NCH2CO2 - ions. When these ions are formed, we once again get a buffer solution in which the pH remains relatively constant until essentially all of the H3N + CH2CO2H molecules have been converted into H2NCH2CO2 - ions.
  • At this point, the pH rises rapidly until it reaches the value observed for a strong base.

The pH titration curve tells us the volume of base required to titrate the positively charged H3N + CH2CO2H molecule to the H3N + CH2CO2 - zwitterion. If we only add half as much base, only half of the positive ions would be titrated to zwitterions. In other words, the concentration of the H3N + CH2CO2H and H3N + CH2CO2 - ions would be the same. Or, using the symbolism in the Henderson-Hasselbach equation:

Because the concentrations of these ions is the same, the logarithm of the ratio of their concentrations is zero.

Thus, at this particular point in the titration curve, the Henderson-Hasselbach equation gives the following equality.

We can therefore determine the pKa of an acid by measuring the pH of a solution in which the acid has been half-titrated.

Because there are two titratable groups in glycine, we get two points at which the amino acid is half-titrated. The first occurs when half of the positive H3N + CH2CO2H molecules have been converted to neutral H3N + CH2CO2 - ions. The second occurs when half of the H3N + CH2CO2 - zwitterions have been converted to negatively charged H2NCH2CO2 - ions.

The following results are obtained when this technique is applied to glycine.

Let's compare these values with the pKa's of acetic acid and the ammonium ion.

The acid/base properties of the a -amino group in an amino acid are very similar to the properties of ammonia and the ammonium ion. The a -amine, however, has a significant effect on the acidity of the carboxylic acid. The -amine increases the value of Ka for the carboxylic acid by a factor of about 100.

The inductive effect of the a -amine can only be felt at the a -CO2H group. If we look at the chemistry of glutamic acid, for example, the a -CO2H group on the R substituent has an acidity that is close to that of acetic acid.

When we titrate an amino acid from the low end of the pH scale (pH 1) to the high end (pH 13), we start with an ion that has a net positive charge and end up with an ion that has a net negative charge.

Somewhere between these extremes, we have to find a situation in which the vast majority of the amino acids are present as the zwitterion with no net electric charge. This point is called the isoelectric point (pI) of the amino acid.

For simple amino acids, in which the R group doesn't contain any titratable groups, the isoelectric point can be calculated by averaging the pKa values for the a -carboxylic acid and a -amino groups. Glycine, for example, has a pI of about 6.

At pH 6, more than 99.98% of the glycine molecules in this solution are present as the neutral H3N + CH2CO2H zwitterion.

When calculating the pI of an amino acid that has a titratable group on the R side chain, it is useful to start by writing the structure of the amino acid at physiological pH (pH 7). Lysine, for example, could be represented by the following diagram.

At physiological pH, lysine has a net positive charge. Thus, we have to increase the pH of the solution to remove positive charge in order to reach the isoelectric point. The pI for lysine is simply the average of the pKa's of the two -NH3 + groups.

At this pH, all of the carboxylic acid groups are present as -CO2 - ions and the total population of the -NH3 + groups is equal to one. Thus, the net charge on the molecule at this pH is zero.

If we apply the same technique to the pKa data for glutamic acid, given above, we get a pI of about 3.1. The three amino acids in this section therefore have very different pI values.

Thus, it isn't surprising that a common technique for separating amino acids (or the proteins they form) involves placing a mixture in the center of a gel and then applying a strong voltage across this gel. This technique, which is known as gel electrophoresis, is based on the fact that amino acids or proteins that carry a net positive charge at the pH at which the separation is done will move toward the negative electrode, whereas those with a net negative charge will move toward the positive electrode.


Important terminologies related with amino acids:

Amino Acids are precursors for proteins, important metabolic intermediates neurotransmitter and precursor for nitrogen containing compounds, glucose, ketone bodies, etc.

Amino acids are organic compound composed of amino (NH2) and acidic (COO-) functional groups.

Amino acids have both amino and carboxylic acid groups attached to Alpha carbon atom have particular importance. There are about 500 naturally occurring amino acids present in nature out of which 20 are standard amino acids commonly found in living organism.

These 20 standard amino acids combines into peptide chain to form the building blocks of proteins. All 20 standard amino acids shows variation in R-chain attached to Alpha carbon.

Sterio-isomerism:

L-isomer is the most common form of Alpha amino acid found naturally. Except glycine Alpha carbon present in all standard amino acids is chiral carbon atom ( attached with 4 different functional groups). Hence all Alpha amino acids can exist in two enantiomers ‘L’ amino acid or ‘D’ amino acid which are nothing but mirror images of each other.

L-amino acids is the most common form in standard amino acids found naturally and in proteins during translation in the ribosome. While D-amino acids founding some proteins produced by post translational modifications after translation with the help of endoplasmic reticulum.

‘D’ form of amino acids are not so common but found in peptidoglycan cell walls of bacteria. It is also found in other compounds like Tyrocidine and Valinomycin.


Interaction of aspartate transcarbamylase with 5-bromocytidine 5'-tri-, di-, and monophosphates

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