[13] Stationary and time-resolved circular dichroism of hemoglobins

Zentz, Christian; Pin, Serge; Alpert, Bernard

doi:10.1016/0076-6879(94)32051-9

Cited by 24 publications

(35 citation statements)

References 89 publications

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“…4). Above 300 nm, the optical activity of the heme results from short and long distance interactions of the heme with the protein matrix, specifically attributed by theoretical calculations to a coupled oscillator interaction between the heme transitions and those of the surrounding aromatic side chains (53,54). This interpretation is consistent with the altered microenvironments of the aromatic amino acids of these ␤6 mutants, as indicated by the fluorescence and UVRR spectroscopic results.…”

Section: Discussionsupporting

confidence: 73%

Solution-active Structural Alterations in Liganded Hemoglobins C (β6 Glu → Lys) and S (β6 Glu → Val)

Hirsch¹,

Juszczak²,

Fataliev³

et al. 1999

Journal of Biological Chemistry

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Based upon existing crystallographic evidence, HbS, HbC, and HbA have essentially the same molecular structure. However, important areas of the molecule are not well defined crystallographically (e.g. the N-terminal nonhelical portion of the ␣ and ␤ chains), and conformational constraints differ in solution and in the crystalline state. Over the years, our laboratory and others have provided evidence of conformational changes in HbS and, more recently, in HbC.We now present data based upon allosteric perturbation monitored by front-face fluorescence, ultraviolet resonance Raman spectroscopy, circular dichroism, and oxygen equilibrium studies that confirm and significantly expand previous findings suggesting solution-active structural differences in liganded forms of HbS and HbC distal to the site of mutation and involving the 2,3-diphosphoglycerate binding pocket. The liganded forms of these hemoglobins are of significant interest because HbC crystallizes in the erythrocyte in the oxy form, and oxy HbS exhibits increased mechanical precipitability and a high propensity to oxidize. Specific findings are as follows: 1) differences in the intrinsic fluorescence indicate that the Trp microenvironments are more hydrophobic for HbS > HbC > HbA, 2) ultraviolet resonance Raman spectroscopy detects alterations in Tyr hydrogen bonding, in Trp hydrophobicity at the ␣ 1 ␤ 2 interface (␤37), and in the A-helix (␣14/␤15) of both chains, 3) displacement by inositol hexaphosphate of the Hb-bound 8-hydroxy-1,3,6-pyrenetrisulfonate (the fluorescent 2,3-diphosphoglycerate analog) follows the order HbA > HbS > HbC, and 4) oxygen equilibria measurements indicate a differential allosteric effect by inositol hexaphosphate for HbC ϳ HbS > HbA.Naturally occurring ␤6 hemoglobin mutants aggregate into defined structures in the erythrocyte. Sickle cell hemoglobin (HbS, ␤6 Glu 3 Val) forms polymers in the deoxy state, whereas HbC (␤6 Glu 3 Lys) forms crystals in the oxy liganded state. A complete understanding of the mechanisms giving rise to deoxy HbS polymers and oxy HbC crystals remains to be elucidated. In the last few years, our laboratory has pursued questions related to mechanisms involved in the ligand-specific, induced crystallization of HbC, starting with site-specific probing of the R-state tetrameric structure of HbC (1, 2). Here, we extend the studies to include a comparison of the R-state of HbS. The R-state of HbS is of particular relevance because oxy HbS exhibits unusual properties compared with HbA, such as mechanical precipitability (3-5), greater unfolding at an airwater interface (6, 7), and increased autooxidation (8 -10).According to existing crystallographic evidence, HbS, HbC, and HbA have essentially the same molecular structure. However, this ignores the following: 1) important areas of the molecule are not well defined crystallographically (e.g. the N-terminal nonhelical portion of the ␣ and ␤ chains), and 2) crystals might constrain conformation compared with that in solution.Spectroscopic and biochemical findin...

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Section: Discussionsupporting

confidence: 73%

Solution-active Structural Alterations in Liganded Hemoglobins C (β6 Glu → Lys) and S (β6 Glu → Val)

Hirsch¹,

Juszczak²,

Fataliev³

et al. 1999

Journal of Biological Chemistry

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“…The two peaks characteristic for this region (Q 0 and Q 1 ), whose relative magnitude gives indications about the constraints at the heme site, 14 reflect symmetric properties of the heme-iron bound material. 15 Also in this region the two proteins show small differences in ellipticity values, probably due to the globins. In this region TBTC induces opposite effects in the two proteins, decreasing ellipticities in the HbIV spectrum from 480 to 580 nm and, on the contrary, increasing the ellipticities of HbI in the whole range (480-620 m).…”

Section: Resultsmentioning

confidence: 93%

“…The 250-300 nm region is indicative of the R-T transition: in the T-form the ellipticity is much more negative than in the corresponding R-form. 15,22 In oxy-HbIV (Fig. 5 A) TBTC reduces the negative value of ellipticity in the region of about 280-400 nm, suggesting a shift towards the R-form and a higher affinity for O 2 .…”

Section: Discussionmentioning

confidence: 93%

“…The region 270-290 nm in various hemoglobins have been used to study changes in the quaternary structure in the hemoglobins at the ␣ 1 ␤ 2 interface. 15,22 In human HbA, the 280-290 nm CD bands should be correlated to the environment of ␣42 and ␤37 aromatic residues, which are maintained in trout HbIV and HbI 23 , but it was related also to the orientation of heme with respect to aromatic amino acids. 15 In this region the spectra obtained for HbIV and HbI are quite different (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…13 The Soret region has been related to the subunit interaction in the ␣ 1 ␤ 1 dimer. 15 Changes in the ␣ 1 ␤ 1 subunit contact should be accompanied by changes in the spatial orientation of aromatic amino acids relative to heme and changes in interaction between heme iron and the proximal histidine. 15 Moreover, possible distortions and/or tilting of the heme macrocycle and hence movements of the particular residues that participate in the rotational strength of the heme transitions have been associated with this region.…”

Section: Discussionmentioning

confidence: 99%

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Steady-state fluorescence and circular dichroism of trout hemoglobins I and IV interacting with tributyltin

et al. 1999

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The effect of tributyltin chloride (TBTC) on rainbow trout (Salmo irideus) hemoglo-bin I (HbI) and hemoglobin IV (HbIV) was characterized by the steady-state fluorescence of intrinsic and extrinsic fluorescent probes. The fluorescence emission spectrum (ex 280 nm) is greatly increased in intensity by the presence of the organotin in both proteins. Circular dichroism spectra in the same samples show a small decrease in 222 , a measure correlated with the percentage of the-helical content. Morever, important changes in near-UV, Soret, and visible regions of CD were induced by TBTC. The correlation of data obtained with trout hemoglo-bins (HbI and HbIV) with similar measurements on globins suggests that the presence of heme is necessary for the interaction of the organotin compound with the proteins. Proteins 1999;34:443-452. 1999 Wiley-Liss, Inc.

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Contribution of ?140Tyr and ?37Trp to the near-UV CD spectra on quaternary structure transition of human hemoglobin A

Nagai

2000

Chirality

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The CD band of human adult hemoglobin (Hb A) at 280 ∼ 290 nm shows a pronounced change from a small positive band to a definite negative band on the oxy (R) to deoxy (T) structural transition. This change has been suggested to be due to environmental alteration of Tyrs (α42, α140, and β145) or β37 Trp residues located at the α1β2 subunit interface by deoxygenation. In order to evaluate contributions of α140Tyr and β37Trp to this change of CD band, we compared the CD spectra of two mutant Hbs, Hb Rouen (α140Tyr→His) and Hb Hirose (β37Trp→Ser) with those of Hb A. Both mutant Hbs gave a distinct, but smaller negative CD band at 287nm in the deoxy form than that of deoxyHb A. Contributions of α140Tyr and β37Trp to the negative band at the 280 ∼ 290 nm region were estimated from difference spectra to be 30% and 26%, respectively. These results indicate that the other aromatic amino acid residues, α42Tyr and β145Tyr, at the α1β2 interface, are also responsible for the change of the CD band upon the R→T transition of Hb A. Chirality 12:216–220, 2000. © 2000 Wiley‐Liss, Inc.

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[13] Stationary and time-resolved circular dichroism of hemoglobins

Cited by 24 publications

References 89 publications

Solution-active Structural Alterations in Liganded Hemoglobins C (β6 Glu → Lys) and S (β6 Glu → Val)

Solution-active Structural Alterations in Liganded Hemoglobins C (β6 Glu → Lys) and S (β6 Glu → Val)

Steady-state fluorescence and circular dichroism of trout hemoglobins I and IV interacting with tributyltin

Contribution of ?140Tyr and ?37Trp to the near-UV CD spectra on quaternary structure transition of human hemoglobin A

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