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A role of heme side-chains of human hemoglobin in its function revealed by circular dichroism and resonance Raman spectroscopy.

ABSTRACT: Structural changes of heme side-chains of human adult hemoglobin (Hb A) upon ligand (O2 or CO) dissociation have been studied by circular dichroism (CD) and resonance Raman (RR) spectroscopies. We point out the occurrence of appreciable deformation of heme side-chains like vinyl and propionate groups prior to the out-of-plane displacement of heme iron. Referring to the recent fine resolved crystal structure of Hb A, the deformations of heme side-chains take place only in the ? subunits. However, these changes are not observed in the isolated ? chain (?4 homotetramer) and, therefore, are associated with the ?-? inter-subunit interactions. For the communications between ? and ? subunits in Hb A regarding signals of ligand dissociation, possible routes are proposed on the basis of the time-resolved absorption, CD, MCD (magnetic CD), and RR spectroscopies. Our finding of the movements of heme side-chains would serve as one of the clues to solve the cooperative O2 binding mechanism of Hb A.

SUBMITTER: Nagai M

PROVIDER: S-EPMC5899723 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

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A role of heme side-chains of human hemoglobin in its function revealed by circular dichroism and resonance Raman spectroscopy.

Nagai Masako M Mizusawa Naoki N Kitagawa Teizo T Nagatomo Shigenori S

Biophysical reviews 20171219 2

Structural changes of heme side-chains of human adult hemoglobin (Hb A) upon ligand (O<sub>2</sub> or CO) dissociation have been studied by circular dichroism (CD) and resonance Raman (RR) spectroscopies. We point out the occurrence of appreciable deformation of heme side-chains like vinyl and propionate groups prior to the out-of-plane displacement of heme iron. Referring to the recent fine resolved crystal structure of Hb A, the deformations of heme side-chains take place only in the β subunit ...[more]

PMID: 29260461

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Project description:BackgroundIn Huntington's disease, expansion of a CAG triplet repeat occurs in exon 1 of the huntingtin gene (HTT), resulting in a protein bearing>35 polyglutamine residues whose N-terminal fragments display a high propensity to misfold and aggregate. Recent data demonstrate that polyglutamine expansion results in conformational changes in the huntingtin protein (HTT), which likely influence its biological and biophysical properties. Developing assays to characterize and measure these conformational changes in isolated proteins and biological samples would advance the testing of novel therapeutic approaches aimed at correcting mutant HTT misfolding. Time-resolved Förster energy transfer (TR-FRET)-based assays represent high-throughput, homogeneous, sensitive immunoassays widely employed for the quantification of proteins of interest. TR-FRET is extremely sensitive to small distances and can therefore provide conformational information based on detection of exposure and relative position of epitopes present on the target protein as recognized by selective antibodies. We have previously reported TR-FRET assays to quantify HTT proteins based on the use of antibodies specific for different amino-terminal HTT epitopes. Here, we investigate the possibility of interrogating HTT protein conformation using these assays.Methodology/principal findingsBy performing TR-FRET measurements on the same samples (purified recombinant proteins or lysates from cells expressing HTT fragments or full length protein) at different temperatures, we have discovered a temperature-dependent, reversible, polyglutamine-dependent conformational change of wild type and expanded mutant HTT proteins. Circular dichroism spectroscopy confirms the temperature and polyglutamine-dependent change in HTT structure, revealing an effect of polyglutamine length and of temperature on the alpha-helical content of the protein.Conclusions/significanceThe temperature- and polyglutamine-dependent effects observed with TR-FRET on HTT proteins represent a simple, scalable, quantitative and sensitive assay to identify genetic and pharmacological modulators of mutant HTT conformation, and potentially to assess the relevance of conformational changes during onset and progression of Huntington's disease.

Dataset Information

A role of heme side-chains of human hemoglobin in its function revealed by circular dichroism and resonance Raman spectroscopy.

Publications

A role of heme side-chains of human hemoglobin in its function revealed by circular dichroism and resonance Raman spectroscopy.

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