Amino Acids

<body> <p align="center"><font size="6"><strong>Amino Acids </strong></font></p> <div align="center"><center><table border="1" width="500"> <tr> <td align="center"><a href="#amino">The Amino Acids</a></td> <td align="center"><a href="#zwitter">Zwitterions</a></td> </tr> <tr> <td align="center"><a href="#table">The Amino Acids Used to Synthesize Proteins</a></td> <td align="center"><a href="#base">The Acid-Base Chemistry of the Amino Acids </a></td> </tr> </table> </center></div><p align="center"><a name="amino"><em><strong>The Amino Acids</strong></em></a></p> <p><strong>Proteins</strong> are formed by polymerizing monomers that are known as <strong>amino acids</strong> because they contain an amine (-NH<sub>2</sub>) and a carboxylic acid (-CO<sub>2</sub>H) 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.</p> <div align="center"><center><table border="0"> <tr> <td><img src="graphics/1.gif" width="83" height="48"></td> </tr> <tr> <td><em>An amino acid</em></td> </tr> </table> </center></div><p align="left">These compounds are known as <font face="Symbol">a</font>-amino acids because the -NH<sub>2</sub> group is on the carbon atom next to the -CO<sub>2</sub>H group, the so-called carbon atom of the carboxylic acid. </p> <p align="center"><a href="#top"><em><strong><img src="graphics/button.gif" alt="return to top" border="0" width="226" height="36"></strong></em></a></p> <p align="center"><a name="zwitter"><em><strong>Zwitterions</strong></em></a></p> <p>The chemistry of amino acids is complicated by the fact that the -NH<sub>2</sub> group is a base and the -CO<sub>2</sub>H group is an acid. In aqueous solution, an H<sup>+</sup> ion is therefore transferred from one end of the molecule to the other to form a <strong>zwitterion</strong> (from the German meaning mongrel ion, or hybrid ion).</p> <p align="center"><img src="graphics/2.gif" width="232" height="49"> </p> <p>Zwitterions are simultaneously electrically charged and electrically neutral. They contain positive and negative charges, but the net charge on the molecule is zero. </p> <p align="center"><a href="#top"><em><strong><img src="graphics/button.gif" alt="return to top" border="0" width="226" height="36"></strong></em></a></p> <p align="center"><a name="table"><em><strong>The Amino Acids Used to Synthesize Proteins</strong></em></a></p> <p>More than 300 amino acids are listed in the <em>Practical Handbook of Biochemistry and Molecular Biology</em>, 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 <em>R</em> substituent. The standard amino acids are therefore classified on the basis of these <em>R</em> groups. Amino acids with nonpolar substituents are said to be <em>hydrophobic</em> (water-hating). Amino acids with polar <em>R</em> groups that form hydrogen bonds to water are classified as <em>hydrophilic</em> (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. </p> <p align="center"><em>The 20 Standard Amino Acids</em></p> <div align="center"><center><table border="0"> <tr> <td valign="top" rowspan="2"><div align="center"><center><table border="0"> <tr> <td width="50"><em>NAME</em></td> <td valign="top"><em>STRUCTURE <br> (AT NEUTRAL pH)</em></td> </tr> <tr> <td colspan="2"><em>Nonpolar (Hydrophobic) R Groups</em></td> </tr> <tr> <td width="50">Glycine (Gly)</td> <td><img src="graphics/1amino.GIF" width="115" height="47"></td> </tr> <tr> <td width="50">Alanine (Ala)</td> <td><img src="graphics/amino2.gif" width="115" height="47"></td> </tr> <tr> <td width="50">Valine (Val)</td> <td><img src="graphics/amino3.gif" width="115" height="69"></td> </tr> <tr> <td width="50">Leucine (Leu)</td> <td><img src="graphics/amino4.gif" width="115" height="99"></td> </tr> <tr> <td width="50">Isoleucine (Ile)</td> <td><img src="graphics/amino5.gif" width="128" height="69"></td> </tr> <tr> <td width="50">Proline (Pro)</td> <td><img src="graphics/amino6.gif" width="119" height="82"></td> </tr> <tr> <td width="50">Methionine (Met)</td> <td><img src="graphics/amino7.gif" width="115" height="137"></td> </tr> <tr> <td width="50">Phenylalanine (Phe)</td> <td><img src="graphics/amino8.gif" width="115" height="135"></td> </tr> <tr> <td width="50">Tryptophan (Trp)</td> <td><img src="graphics/amino9.gif" width="142" height="164"></td> </tr> <tr> <td colspan="2"><em>Polar (Hydrophilic) R Groups</em></td> </tr> <tr> <td>Serine<br> (ser)</td> <td><img src="graphics/amino10.gif" width="115" height="47"></td> </tr> <tr> <td>Threonine<br> (Thr)</td> <td><img src="graphics/amino11.gif" width="115" height="69"></td> </tr> <tr> <td>Tyrosine<br> (Tyr)</td> <td><img src="graphics/amino12.gif" width="115" height="167"></td> </tr> <tr> <td>Cysteine<br> (Cys)</td> <td><img src="graphics/amino13.gif" width="115" height="47"></td> </tr> <tr> <td>Asparagine<br> (Asn)</td> <td><img src="graphics/amino14.gif" width="115" height="114"></td> </tr> <tr> <td>Glutamine<br> (Gln)</td> <td><img src="graphics/amino15.gif" width="115" height="137"></td> </tr> <tr> <td colspan="2"><em>Negatively Charged R Groups</em></td> </tr> <tr> <td>Aspartic acid (Asp)</td> <td><img src="graphics/amino16.gif" width="115" height="77"></td> </tr> <tr> <td>Glutamic acid <br> (Glu)</td> <td><img src="graphics/amino17.gif" width="115" height="107"></td> </tr> <tr> <td colspan="2"><em>Positively Charged R Groups</em></td> </tr> <tr> <td>Lysine<br> (Lys)</td> <td><img src="graphics/amino18.gif" width="115" height="170"></td> </tr> <tr> <td>Arginine<br> (Arg)</td> <td><img src="graphics/amino19.gif" width="120" height="197"></td> </tr> <tr> <td>Histidine<br> (His)</td> <td><img src="graphics/amino20.gif" width="144" height="128"></td> </tr> </table> </center></div></td> </tr> </table> </center></div><p align="center"> </p> <div align="center"><center><table border="5" cellpadding="10" bordercolor="#B87333"> <tr> <td><a name="prob1"><em><strong>Practice Problem 1:</strong></em></a><p>Use the structures of the following amino acids in the table of <a href="#table">standard amino acids </a>to classify these compounds as either nonpolar/hydrophobic, polar/hydrophilic, negatively charged/hydrophilic, or positively charged/hydrophilic. </p> <p>(a) Valine: <em>R</em> = -CH(CH<sub>3</sub>)<sub>2</sub> </p> <p>(b) Serine: <em>R</em> = -CH<sub>2</sub>OH </p> <p>(c) Aspartic acid: <em>R</em> = -CH<sub>2</sub>CO<sub>2</sub><sup>-</sup> </p> <p>(d) Lysine: <em>R</em> = -(CH<sub>2</sub>)<sub>4</sub>NH<sub>3</sub><sup>+</sup> </p> <p><a href="prob1.html"><em><strong>Click here to check your answer to Practice Problem 1</strong></em></a></p> </td> </tr> </table> </center></div><p align="center"><a href="#top"><em><strong><img src="graphics/button.gif" alt="return to top" border="0" width="226" height="36"></strong></em></a></p> <p align="center"><em><strong>Amino Acids as Stereoisomers</strong></em></p> <p>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. </p> <div align="center"><center><table border="0"> <tr> <td><img src="graphics/3.gif" width="95" height="77"></td> <td> </td> <td><img src="graphics/4.gif" width="91" height="77"></td> </tr> <tr> <td align="center"><em>D-Alanine</em></td> <td align="center"> </td> <td align="center"><em>L-Alanine</em></td> </tr> </table> </center></div><p>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<em><strong><img src="graphics/em.gif" alt="--" width="22" height="9"> </strong></em>a decarboxylated derivative of histidine <em><strong><img src="graphics/em.gif" alt="--" width="22" height="9"> </strong></em>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 <em>Awakening</em>, 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. </p> <div align="center"><center><table border="0"> <tr> <td><img src="graphics/5.gif" width="159" height="90"></td> <td><em>Histamine</em></td> <td> </td> <td><em><img src="graphics/6.gif" width="216" height="90"></em></td> <td><em>L-DOPA</em></td> </tr> <tr> <td> </td> <td> </td> <td> </td> <td> </td> <td> </td> </tr> <tr> <td colspan="2"><img src="graphics/7.gif" width="336" height="118"></td> <td> </td> <td><em>Thyroxine</em></td> <td> </td> </tr> </table> </center></div><p align="center"><a href="#top"><img src="graphics/button.gif" alt="return to top" border="0" width="226" height="36"></a></p> <p align="center"><a name="base"><em><strong>The Acid-Base Chemistry of the Amino Acids </strong></em></a></p> <p>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. </p> <p align="center"><img src="graphics/img6.gif" width="405" height="70"> </p> <p>Thus, it is not surprising that an H<sup>+</sup> ion is transferred from one end of the molecule to the other when an amino acid dissolves in water. </p> <p align="center"><img src="graphics/8.gif" width="242" height="50"> </p> <p>The zwitterion is the dominant species in aqueous solutions at physiological pH (pH <img src="graphics/congrnt.gif" width="20" height="11"> 7). The zwitterion can undergo acid-base reactions, howeer, if we add either a strong acid or a strong base to the solution. </p> <p>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 -CO<sub>2</sub><sup>-</sup> end of this molecule picks up an H<sup>+</sup> ion to form a molecule with a net positive charge. </p> <p align="center"><img src="graphics/9.gif" width="238" height="49"></p> <p align="left">In the presence of a strong base, the -NH<sub>3</sub><sup>+</sup> end of the molecule loses an H<sup>+</sup> ion to form a molecule with a net negative charge. </p> <p align="center"><img src="graphics/10.gif" width="225" height="49"> </p> <p><a name="curve"></a>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.</p> <p align="center"><img src="graphics/21.gif" width="441" height="332"> </p> <p>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. </p> <p align="center"><img src="graphics/img10.gif" width="555" height="63"> </p> <p>Let's now rearrange the <em>K</em><sub>a</sub> expression, </p> <p align="center"><img src="graphics/img11.gif" width="555" height="59"> </p> <p align="left">take the log to the base 10 of both sides of this equation, </p> <p align="center"><img src="graphics/img12.gif" width="555" height="59"> </p> <p>and then multiply both sides of the equation by -1. </p> <p align="center"><img src="graphics/img13.gif" width="555" height="59"> </p> <p>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 p<em>K</em><sub>a</sub> of the acid. </p> <p align="center"><img src="graphics/img14.gif" width="555" height="59"> </p> <p>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 </p> <p align="center"><img src="graphics/img15.gif" width="555" height="62"> </p> <p>to give the following form of this equation. </p> <p align="center"><img src="graphics/img16.gif" width="555" height="59"> </p> <p>This equation is known as the <strong>Henderson-Hasselbach equation</strong>, and it can be used to calculate the pH of the solution at any point in the <a href="#curve">titration curve. </a></p> <p>The following occurs as we go from left to right across this titration curve. </p> <ul> <li>The pH initially increases as we add base to the solution because the base deprotonates some of the positively charged H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub>H ions that were present in the strongly acidic solution. </li> <li>The pH then levels off because we form a buffer solution in which we have reasonable concentrations of both an acid, H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub>H, and its conjugate base, H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub><sup>-</sup>. </li> <li>When virtually all of the H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub>H molecules have been deprotonated, we no longer have a buffer solution and the pH rises rapidly when more NaOH is added to the solution.</li> <li>The pH then levels off as some of the neutral H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub><sup>-</sup> molecules lose protons to form negatively charged H<sub>2</sub>NCH<sub>2</sub>CO<sub>2</sub><sup>-</sup> 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 H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub>H molecules have been converted into H<sub>2</sub>NCH<sub>2</sub>CO<sub>2</sub><sup>-</sup> ions.</li> <li>At this point, the pH rises rapidly until it reaches the value observed for a strong base.</li> </ul> <p>The pH titration curve tells us the volume of base required to titrate the positively charged H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub>H molecule to the H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub><sup>-</sup> 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 H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub>H and H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub><sup>-</sup> ions would be the same. Or, using the symbolism in the Henderson-Hasselbach equation: </p> <p align="center">[HA] = [A<sup>-</sup>] </p> <p>Because the concentrations of these ions is the same, the logarithm of the ratio of their concentrations is zero. </p> <p align="center"><img src="graphics/img17.gif" width="555" height="59"> </p> <p>Thus, at this particular point in the titration curve, the Henderson-Hasselbach equation gives the following equality. </p> <p align="center">pH = p<em>K</em><sub>a</sub> </p> <p>We can therefore determine the p<em>K</em><sub>a</sub> of an acid by measuring the pH of a solution in which the acid has been half-titrated. </p> <p>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 H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub>H molecules have been converted to neutral H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub><sup>-</sup> ions. The second occurs when half of the H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub><sup>-</sup> zwitterions have been converted to negatively charged H<sub>2</sub>NCH<sub>2</sub>CO<sub>2</sub><sup>-</sup> ions. </p> <p>The following results are obtained when this technique is applied to glycine. </p> <p align="center"><img src="graphics/img18.gif" width="174" height="61"> </p> <p>Let's compare these values with the pK<sub>a</sub>'s of acetic acid and the ammonium ion. </p> <div align="center"><center><table border="0"> <tr> <td>CH<sub>3</sub>CO<sub>2</sub>H</td> <td> </td> <td> </td> <td>p<em>K</em><sub>a</sub> = 4.74 </td> <td> </td> </tr> <tr> <td>NH<sub>4</sub><sup>+</sup></td> <td> </td> <td> </td> <td>p<em>K</em><sub>a</sub> = 9.24 </td> <td> </td> </tr> </table> </center></div><p>The acid/base properties of the <font face="Symbol">a</font>-amino group in an amino acid are very similar to the properties of ammonia and the ammonium ion. The <font face="Symbol">a</font>-amine, however, has a significant effect on the acidity of the carboxylic acid. The -amine increases the value of <em>K</em><sub>a</sub> for the carboxylic acid by a factor of about 100. </p> <p>The inductive effect of the <font face="Symbol">a</font>-amine can only be felt at the <font face="Symbol">a</font>-CO<sub>2</sub>H group. If we look at the chemistry of glutamic acid, for example, the <font face="Symbol">a</font>-CO<sub>2</sub>H group on the <em>R</em> substituent has an acidity that is close to that of acetic acid. </p> <p align="center"><img src="graphics/11.gif" width="172" height="153"> </p> <p>When we titrate an amino acid from the low end of the pH scale (pH <img src="graphics/congrnt.gif" width="20" height="11"> 1) to the high end (pH <img src="graphics/congrnt.gif" width="20" height="11">13), we start with an ion that has a net positive charge and end up with an ion that has a net negative charge. </p> <p align="center"><img src="graphics/12.gif" width="252" height="47"> </p> <p>Somewhere between these extremes, we have to find a situation in which the vast majority of the amino acids are present as the zwitterion <em><strong><img src="graphics/em.gif" alt="--" width="22" height="9"></strong></em> with no net electric charge. This point is called the <em><strong>isoelectric point (pI)</strong></em> of the amino acid. </p> <p>For simple amino acids, in which the <em>R</em> group doesn't contain any titratable groups, the isoelectric point can be calculated by averaging the <em>pK</em><sub><em>a</em></sub> values for the <font face="Symbol">a</font>-carboxylic acid and <font face="Symbol">a</font>-amino groups. Glycine, for example, has a p<em>I</em> of about 6. </p> <p align="center">p<em>I</em> = <u>2.35 + 9.78</u> = 6.1 <br> 2 </p> <p>At pH <img src="graphics/congrnt.gif" width="20" height="11"> 6, more than 99.98% of the glycine molecules in this solution are present as the neutral H<sub>3</sub>N<sup>+</sup>CH<sub>2</sub>CO<sub>2</sub>H zwitterion. </p> <p>When calculating the p<em>I</em> of an amino acid that has a titratable group on the <em>R</em> side chain, it is useful to start by writing the structure of the amino acid at physiological pH (pH <img src="graphics/congrnt.gif" width="20" height="11"> 7). Lysine, for example, could be represented by the following diagram. </p> <p align="center"><img src="graphics/img21.gif" align="middle" width="191" height="130"> </p> <p>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 p<em>I</em> for lysine is simply the average of the pK<sub>a</sub>'s of the two -NH<sub>3</sub><sup>+</sup> groups. </p> <p align="center">p<em>I</em> = <u>9.18 + 10.79</u> <img src="graphics/congrnt.gif" width="20" height="11"> 10.0 <br> 2 </p> <p>At this pH, all of the carboxylic acid groups are present as -CO<sub>2</sub><sup>-</sup> ions and the total population of the -NH<sub>3</sub><sup>+</sup> groups is equal to one. Thus, the net charge on the molecule at this pH is zero. </p> <p>If we apply the same technique to the p<em>K</em><sub>a</sub> data for glutamic acid, given above, we get a p<em>I</em> of about 3.1. The three amino acids in this section therefore have very different p<em>I</em> values. </p> <div align="center"><center><table border="0"> <tr> <td>Glutamic acid </td> <td> </td> <td>(<em>R</em> = -CH<sub>2</sub>CH<sub>2</sub>CO<sub>2</sub><sup>-</sup>): </td> <td> </td> <td>p<em>I</em> = 3.1 </td> </tr> <tr> <td>Glycine </td> <td> </td> <td>(<em>R</em> = -H):</td> <td> </td> <td>p<em>I</em> = 6.1 </td> </tr> <tr> <td>Lysine </td> <td> </td> <td>(<em>R</em> = -CH<sub>2</sub>CH<sub>2</sub>CH<sub>2</sub>CH<sub>2</sub>NH<sub>3</sub><sup>+</sup>):</td> <td> </td> <td>p<em>I</em> = 10.0</td> </tr> </table> </center></div><p>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 <strong>gel electrophoresis</strong>, 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. </p> <p align="center"><a href="#top"><img src="graphics/button.gif" alt="return to top" border="0" width="226" height="36"></a></p> <hr> <p><img src="http://chemed.chem.purdue.edu/Count.cgi?df=toprev_bio_amino2.dat"></p> </body>