Histidine (His, H) (Molecular Biology)

The amino acid histidine is incorporated into the nascent polypeptide chain during protein biosynthesis in response to two codons—CAU and CAC—and represents approximately 2.2% of the residues of the proteins that have been characterized. The histidyl residue incorporated has a mass of 137.14 Da, a van der Waals volume of 118 A 3, and an accessible surface area of 194 A2. His residues are changed during divergent evolution with average frequency; they are interchanged in homologous proteins most frequently with asparagine and glutamine residues.

The side chain of His residues consists of a b-methylene carbon and an imidazole group:

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The imidazole groups possesses several special properties that make it extremely effective as a nucleophilic catalyst. It is an amine, which is much more reactive than hydroxide ion in terms of basicity. Furthermore, it is a tertiary amine, which is intrinsically more nucleophilic than primary or secondary amines. The enhanced reactivity of tertiary amines is usually canceled by their greater steric hindrance, but in imidazole the atoms bonded to the two nitrogen atoms are held back in a five-membered ring and cause relatively little steric hindrance. Imidazole has a pKa value near 7, so


it is one of the strongest bases that can exist at neutral pH. A weaker base would have a lower nucleophilic reactivity, whereas a stronger base would be protonated to a greater extent at neutral pH and would be correspondingly less reactive.

In the nonionized form of the imidazole ring, the nitrogen atom with the hydrogen atom is an electrophile and a donor for hydrogen bonding, while the other nitrogen atom is a nucleophile and a hydrogen bond acceptor. Consequently, this one side chain is extremely versatile, almost the chemical equivalent of being ambidextrous. Not surprisingly, His residues are often found at the active sites of enzymes and involved directly in catalysis.

The two nitrogen atoms of the His side chain are designated here as dl and e 2, but they are also known, respectively, as p and q or as N-1 and N-3. The latter designation is often ambiguous, because biochemists usually assign the number 1 to the nitrogen atom adjacent to the Cg, whereas organic chemists tend to reverse the numbers. The nonionized imidazole ring can exist as two tautomers, with the hydrogen atom on either the dl and 6 2 nitrogen atom. In model peptides, the hydrogen atom is usually predominantly on the e 2 nitrogen atom, which has a pK a value about 0.6 pH unit higher than that of the d1 atom. The relative affinities of the two nitrogen atoms for protons can vary with their local environment, however, and both forms are found in folded proteins.

The nonionized His side chain is readily protonated, with a p Ka value near 7 at the second N atom, which destroys its nucleophilicity. The positive charge is shared by the two N atoms by resonance:

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The Cf 1 atom, between the two N atoms, is observed to exchange its hydrogen atom slowly with aqueous solvents, indicating that it has a very small probability of being deprotonated:

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This hydrogen exchange reaction provides a useful probe of the environments of His residues in proteins. His residues are especially useful in 1H-NMR studies of proteins because the hydrogen atom on the CP1 atom is usually well resolved from the multitude of resonances of the other hydrogen atoms in proteins. Its resonance is usually shifted by about 1 ppm to a lower field strength on protonation of the side chain, often making it relatively easy to determine the pKa values of individual His residues, even in large proteins.

His is one of the residues least favoring the alpha-helical conformation in model peptides, but it occurs in a-helices in native protein structures slightly more frequently than in beta -sheets or reverse turns .

The imidazole group is, in principle, capable of undergoing numerous chemical modification reactions, but most of these occur much more readily with amino and thiol groups, so very few are suitable for modifying His residues specifically. The imidazole nitrogen atoms have an intrinsic affinity for metal ions, especially zinc, iron, and copper. His residues are frequently involved in binding such ligands to proteins, as in metal-requiring enzymes, zinc fingers, myoglobin, and hemoglobin.

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