34. Haemoglobin has characteristic circular dichroism (CD) peaks in the far-UV, near UV and Soret regions. Contribution to near-UV CD comes entirely from

(A) Aromatic amino acid residues.

(B) Heme group.

(C) Heme and aromatic amino acid residues.

(D) Peptide bonds and aromatic amino acid residues.

Near-UV Circular Dichroism of Hemoglobin: Contribution of Aromatic Amino Acid Residues

Correct Answer

Option (1): Aromatic amino acid residues.

Explanation

Circular Dichroism (CD) spectroscopy is widely used to investigate the structure of proteins because different regions of the ultraviolet and visible spectrum provide information about different structural components of a protein molecule. In haemoglobin, three distinct spectral regions are commonly analyzed: the far-UV region, the near-UV region, and the Soret region. Each region originates from different chromophores and therefore provides different structural information.

The near-UV region, which extends approximately from 250 to 320 nm, is dominated entirely by the electronic transitions of the aromatic amino acid side chains. The principal contributors are tryptophan, tyrosine, and phenylalanine. These aromatic residues generate CD signals only when they are present in an asymmetric or chiral environment within the folded protein. Consequently, the near-UV CD spectrum reflects the tertiary structure of the protein and provides valuable information about the spatial arrangement and packing of aromatic side chains.

In haemoglobin, although the heme prosthetic group is an essential structural component, it does not contribute to the near-UV CD spectrum. Instead, the heme group produces characteristic CD signals in the Soret region, where its intense electronic absorption band is located. Therefore, the near-UV CD signal arises exclusively from the aromatic amino acid residues present in the globin chains.

Why Option (1) is Correct

The near-UV CD spectrum originates from the aromatic side chains of tryptophan, tyrosine, and phenylalanine. These residues become optically active because they are located in a chiral three-dimensional environment created by protein folding. Since the question specifically asks about the contribution to the near-UV CD region, aromatic amino acid residues are the only correct answer.

Why Option (2) is Incorrect

The heme group is responsible for the characteristic CD signals observed in the Soret region, generally around 400–450 nm. It does not contribute to the near-UV CD spectrum. Therefore, attributing the near-UV signal solely to the heme group is incorrect.

Why Option (3) is Incorrect

This option incorrectly includes the heme group as a contributor to the near-UV CD spectrum. While haemoglobin contains both aromatic amino acids and heme, their spectroscopic contributions occur in different wavelength regions. Aromatic amino acid residues dominate the near-UV region, whereas the heme group contributes predominantly to the Soret region.

Why Option (4) is Incorrect

Peptide bonds contribute mainly to the far-UV CD spectrum, typically between 190 and 250 nm, where transitions associated with the peptide backbone are observed. The near-UV region does not receive significant contributions from peptide bonds. Therefore, this option incorrectly combines chromophores that belong to different spectral regions.

Contribution of Different Regions in Circular Dichroism Spectroscopy

The far-UV region (approximately 190–250 nm) primarily arises from the peptide backbone and is used to determine secondary structural elements such as α-helices, β-sheets, and random coils. The near-UV region (250–320 nm) originates from aromatic amino acid residues and provides information about the tertiary structure of proteins. The Soret region (approximately 380–450 nm) is characteristic of heme-containing proteins and reflects the electronic transitions of the heme prosthetic group and its interactions with the surrounding protein environment.

Importance of the Near-UV CD Spectrum

The near-UV CD spectrum is highly sensitive to the three-dimensional arrangement of aromatic amino acid residues. Changes in protein folding, ligand binding, mutations, or conformational rearrangements alter the environment surrounding these residues, producing measurable changes in the CD spectrum. As a result, near-UV CD spectroscopy is particularly useful for monitoring tertiary structural changes while preserving the protein in its native solution state.

Conclusion

The near-UV Circular Dichroism spectrum of haemoglobin is produced exclusively by the aromatic amino acid residues present within the protein. The peptide backbone contributes to the far-UV region, whereas the heme prosthetic group contributes to the Soret region. Therefore, the correct answer is Option (1): Aromatic amino acid residues.