Identifying Amino Acids - Biochemistry

Agarose gel electrophoresis is typically used for large molecules, not small ones like individual amino acids. Isoelectric focusing separates molecules by isoelectric point. Edman degradation only permits sequencing of about 30 amino acids, not an unlimited amount. High-performance liquid chromatography separates proteins which are peptide sequences.

Most amino acids that exist in living organisms contain an alpha-carbon that is a stereocenter (or chiral center). This means that this central carbon is bonded to four different substituents, thus allowing the amino acid to have two possible isomers. Although practically all amino acids in nature occur as the L-isomer, glycine is an exception. The reason for this is that glycine's alpha-carbon is bonded to two hydrogen atoms. As a result, this alpha-carbon is not a stereocenter, which means that glycine only has one possible conformation with no isomers.

Tryptophan contains a five membered ring and a six membered ring on its side group. Tyrosine and phenylalanine both contain one six membered ring. Histidine contains a five membered ring on its side chain. Threonine's R-group is , and does not have a ring structure.

The question states that monoamines are synthesized from aromatic amino acids. Aromatic amino acids are defined as those that have one or more aromatic rings in their structure. There are three main aromatic amino acids: histidine, tryptophan and tyrosine. Tyrosine is the precursor for many of the catecholamines (such as norepinephrine, epinephrine and dopamine) whereas histidine is the precursor for histamine. Tryptophan is the precursor for tyrosine; therefore, a lack of tryptophan will result in decreased catecholamine synthesis.

At a pH of 12, the amino group for all of the amino acids would be deprotonated, resulting in at least a charge from the acid group. Another negative charge comes from the carboxyl group on the backbone. Tyrosine is amphipathic, and the pKa of its sidechain is 10.8, meaning it is a deprotonated at a pH of 12. This gives it a charge at a pH of 12. Tryptophan has no side-chain pKa so 3 equivalence points would not be seen. Threonine would also not have 3 equivalence points. Glutamic acid doesn't have amphipathic properties. Histidine is amphipathic, but it is a basic amino acid.

All amino acids have an amino group, a carboxyl group, a hydrogen, and an R-group that is unique to the amino acid. In this structure, the R-group is a hydrogen, which corresponds to the amino acid glycine.

Since this is an ion-exchange chromatography method, we expect that the protein found in the column has the opposite charge of the beads. Since the beads were negatively charged, we expect the amino acid to be positively charged. Lysine has a basic side chain that can easily pick up a hydrogen from solution and become positively charged.

Only aspartate and threonine have side chains capable of hydrogen bonding interactions. Aspartate has a terminal carboxylate which can act as a hydrogen bond acceptor and as a hydrogen bond donor when protonated. Threonine has a terminal hydroxyl group which can also act as a hydrogen bond donor. Alanine has an entirely aliphatic side chain which is unable to participate in hydrogen bonding, and methionine has a sulfhydryl group, that cannot participate in hydrogen bonding.

We're told in the question stem that a mutation is causing a single amino acid substitution. Originally, the amino acid encoded by this region was a lysine residue. To find the amino acid that would cause the least detriment to the organism, we need to recognize which amino acid is the most similar to lysine. Since lysine is a basic amino acid under physiological conditions, it will tend to carry a positive charge most of the time. Therefore, we are looking for an amino acid that is also basic and carries a positive charge under physiological conditions. Arginine is one such amino acid that meets this criteria, and thus it is the correct answer. Glutamate is incorrect because it is acidic under physiological conditions, and thus will carry a negative charge. Glycine, valine, and tryptophan are also all incorrect because each of these amino acids are neutral under physiological conditions.

We're told in the question that this amino acid has a pI = 10.4. Therefore, we expect this to be a basic amino acid. This means that one carboxyl group, one amino group, and one basic R-group will be present.

At the start of the titration, the solution starts out acidic at a very low pH. At a pH of 2, since so many protons are in solution at this pH, all of the important functional groups under consideration will be protonated. Thus, the amino acid will have a positive charge on each of its basic functional groups, and a neutral charge on its carboxyl group, giving the amino acid a net charge of +2.

Once the pH has climbed to a value of 8, we would expect that only the carboxyl group will be deprotonated. As a result, the carboxyl group will have a negative charge, while each of the other two basic functional groups will still retain their positive charge. Thus, the amino acid at this pH will have a net charge of +1.

And finally, once the pH climbs all the way up to a value of 12, we can expect the two basic functional groups to be deprotonated. Consequently, each of the basic functional groups will be neutral, while the carboxyl group will still be negative. Thus, the overall charge of the amino acid will be negative at this pH.

So overall, the amino acid will be positive, then positive, followed by negative.

Because proline has part of its R-group side chain attached to its amino group, it has a unique structure that differs from all other amino acids. The ring structure that it forms with its amino groups causes it to be unable to be found in alpha helix protein structures - if it were to be in one the structure would be disrupted. However, proline is important in kinks and turns because of its small, unique structure.

This problem tests knowledge of amino acid pKa values. Because the R-group of glutamic acid has a very low pKa of 4.25, which is less than the solution pH of 9.0, it will be negatively charged. Lysine and arginine are both positively charged at this pH, whereas tyrosine and methionine are both neutral.

The guanidino group of arginine is protonated in solutions with pH at or below the pKa of 12.5. It is a positively charged basic side chain that becomes neutral at a pH 12.5 or greater.

Serine and tyrosine both have uncharged polar side chains with one hydroxyl group, but only tyrosine has a pKa value for its side chain of 10.5. This indicates that tyrosine will lose its proton above pH 10.5 and is therefore neutral at pH 7.0.

All the amino acids (except glycine) are chiral, but threonine and isoleucine are the only two with a second asymmetric carbon. As a result, they give two pairs of enantiomers that have different physical properties.

Phenylalanine (F), tyrosine (Y), and tryptophan (W) all contain bulky aromatic side groups that cause proteins to absorb UV light at 280 nm.

Amino acids can be identified by their full names, three-letter codes, and one-letter symbols. Glutamine has the three-letter code "gln" and the one-letter symbol "Q." Glutamic acid has the three-letter code "glu" and the one-letter symbol "E." Lysine has the three-letter code "lys" and the one-letter symbol "K." Aspartic acid has the three-letter code "asp" and the one-letter symbol "D."

The structure of this amino acid classifies it as histidine.

Considering that the protein was run through a cation exchange column, one would expect that the more positively charged proteins would flow very slowly through the column. A cation exchange column is coated with negatively charged beads that would become attracted to the positively charged elements of the protein, causing them to elute out of the column slowly. Lysine and Arginine are positively charged amino acids that would cause this to happen, and a protein rich in these amino acids would elute slowly when run through a cation exchange column.

The question states that the amino acid is placed in an acidic solution. This means that the amino groups and the carboxylic acid groups on the amino acids will be protonated (due to the excess hydrogen ions found in an acidic environment). When they become protonated, basic groups like amines become positively charged whereas acidic groups like carboxylic acids become neutral. Basic amino acids have extra basic groups (like amino groups) whereas acidic amino acids have extra acidic groups (like carboxylic acids). Of the listed amino acids, arginine and lysine are basic amino acids and will have a positive charge (due to the protonated basic groups). There are three basic amino acids (arginine, lysine and histidine) whereas there two acidic amino acids (aspartic acid and glutamic acid). Leucine is neither acidic nor basic.