Expression and properties of recombinant HbA2 (α2δ2) and hybrids containing δ-β sequences

Kenji Inagaki, Junko Inagaki, A. Dumoulin, J. C. Padovan, B. T. Chait, A. Popowicz, L. R. Manning, J. M. Manning

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Hemoglobin A22δ2), which is present at low concentration (1-2%) in the circulating red cells of normal individuals, has two important features that merit its study, i.e., it inhibits polymerization of sickle HbS and its elevated concentration in some thalassemias is a useful clinical diagnostic. However, reports on its functional properties regarding O2 binding are conflicting. We have attempted to resolve these discrepancies by expressing, for the first time, recombinant hemoglobin A2 and systematically studying its functional properties. The construct expressing HbA2 contains only α and δ genes so that the extensive purification required to isolate natural HbA2 is circumvented. Although natural hemoglobin A2 is expressed at low levels in vivo, the amount of recombinant α2δ2 expressed in yeast is similar to that found for adult hemoglobin A and for fetal hemoglobin F when the α + β or the α + γ genes, respectively, are present on the construct. Recombinant HbA2 is stable, i.e., not easily oxidized, and it is a cooperative functional hemoglobin with tetramer-dimer dissociation properties like those of adult HbA. However, its intrinsic oxygen affinity and response to the allosteric regulators chloride and 2,3-diphosphoglycerate are lower than the corresponding properties for adult hemoglobin. Molecular modeling studies which attempt to understand these properties of HbA2 are described.

Original languageEnglish
Pages (from-to)649-662
Number of pages14
JournalJournal of Protein Chemistry
Volume19
Issue number8
DOIs
Publication statusPublished - 2000

Fingerprint

Hemoglobin A2
Hemoglobin
Fetal Hemoglobin
Hemoglobins
Genes
Hemoglobin A
2,3-Diphosphoglycerate
Molecular modeling
Thalassemia
Polymerization
Dimers
Yeast
Purification
Chlorides
Yeasts
Cells
Oxygen

Keywords

  • Allosteric regulator
  • Delta gene
  • Hemoglobin
  • Tetramer strength

ASJC Scopus subject areas

  • Biochemistry

Cite this

Expression and properties of recombinant HbA2 (α2δ2) and hybrids containing δ-β sequences. / Inagaki, Kenji; Inagaki, Junko; Dumoulin, A.; Padovan, J. C.; Chait, B. T.; Popowicz, A.; Manning, L. R.; Manning, J. M.

In: Journal of Protein Chemistry, Vol. 19, No. 8, 2000, p. 649-662.

Research output: Contribution to journalArticle

Inagaki, Kenji ; Inagaki, Junko ; Dumoulin, A. ; Padovan, J. C. ; Chait, B. T. ; Popowicz, A. ; Manning, L. R. ; Manning, J. M. / Expression and properties of recombinant HbA2 (α2δ2) and hybrids containing δ-β sequences. In: Journal of Protein Chemistry. 2000 ; Vol. 19, No. 8. pp. 649-662.
@article{9435a98fc2ab4e39956e256b990537b0,
title = "Expression and properties of recombinant HbA2 (α2δ2) and hybrids containing δ-β sequences",
abstract = "Hemoglobin A2 (α2δ2), which is present at low concentration (1-2{\%}) in the circulating red cells of normal individuals, has two important features that merit its study, i.e., it inhibits polymerization of sickle HbS and its elevated concentration in some thalassemias is a useful clinical diagnostic. However, reports on its functional properties regarding O2 binding are conflicting. We have attempted to resolve these discrepancies by expressing, for the first time, recombinant hemoglobin A2 and systematically studying its functional properties. The construct expressing HbA2 contains only α and δ genes so that the extensive purification required to isolate natural HbA2 is circumvented. Although natural hemoglobin A2 is expressed at low levels in vivo, the amount of recombinant α2δ2 expressed in yeast is similar to that found for adult hemoglobin A and for fetal hemoglobin F when the α + β or the α + γ genes, respectively, are present on the construct. Recombinant HbA2 is stable, i.e., not easily oxidized, and it is a cooperative functional hemoglobin with tetramer-dimer dissociation properties like those of adult HbA. However, its intrinsic oxygen affinity and response to the allosteric regulators chloride and 2,3-diphosphoglycerate are lower than the corresponding properties for adult hemoglobin. Molecular modeling studies which attempt to understand these properties of HbA2 are described.",
keywords = "Allosteric regulator, Delta gene, Hemoglobin, Tetramer strength",
author = "Kenji Inagaki and Junko Inagaki and A. Dumoulin and Padovan, {J. C.} and Chait, {B. T.} and A. Popowicz and Manning, {L. R.} and Manning, {J. M.}",
year = "2000",
doi = "10.1023/A:1007196118200",
language = "English",
volume = "19",
pages = "649--662",
journal = "Protein Journal",
issn = "1572-3887",
publisher = "Springer New York",
number = "8",

}

TY - JOUR

T1 - Expression and properties of recombinant HbA2 (α2δ2) and hybrids containing δ-β sequences

AU - Inagaki, Kenji

AU - Inagaki, Junko

AU - Dumoulin, A.

AU - Padovan, J. C.

AU - Chait, B. T.

AU - Popowicz, A.

AU - Manning, L. R.

AU - Manning, J. M.

PY - 2000

Y1 - 2000

N2 - Hemoglobin A2 (α2δ2), which is present at low concentration (1-2%) in the circulating red cells of normal individuals, has two important features that merit its study, i.e., it inhibits polymerization of sickle HbS and its elevated concentration in some thalassemias is a useful clinical diagnostic. However, reports on its functional properties regarding O2 binding are conflicting. We have attempted to resolve these discrepancies by expressing, for the first time, recombinant hemoglobin A2 and systematically studying its functional properties. The construct expressing HbA2 contains only α and δ genes so that the extensive purification required to isolate natural HbA2 is circumvented. Although natural hemoglobin A2 is expressed at low levels in vivo, the amount of recombinant α2δ2 expressed in yeast is similar to that found for adult hemoglobin A and for fetal hemoglobin F when the α + β or the α + γ genes, respectively, are present on the construct. Recombinant HbA2 is stable, i.e., not easily oxidized, and it is a cooperative functional hemoglobin with tetramer-dimer dissociation properties like those of adult HbA. However, its intrinsic oxygen affinity and response to the allosteric regulators chloride and 2,3-diphosphoglycerate are lower than the corresponding properties for adult hemoglobin. Molecular modeling studies which attempt to understand these properties of HbA2 are described.

AB - Hemoglobin A2 (α2δ2), which is present at low concentration (1-2%) in the circulating red cells of normal individuals, has two important features that merit its study, i.e., it inhibits polymerization of sickle HbS and its elevated concentration in some thalassemias is a useful clinical diagnostic. However, reports on its functional properties regarding O2 binding are conflicting. We have attempted to resolve these discrepancies by expressing, for the first time, recombinant hemoglobin A2 and systematically studying its functional properties. The construct expressing HbA2 contains only α and δ genes so that the extensive purification required to isolate natural HbA2 is circumvented. Although natural hemoglobin A2 is expressed at low levels in vivo, the amount of recombinant α2δ2 expressed in yeast is similar to that found for adult hemoglobin A and for fetal hemoglobin F when the α + β or the α + γ genes, respectively, are present on the construct. Recombinant HbA2 is stable, i.e., not easily oxidized, and it is a cooperative functional hemoglobin with tetramer-dimer dissociation properties like those of adult HbA. However, its intrinsic oxygen affinity and response to the allosteric regulators chloride and 2,3-diphosphoglycerate are lower than the corresponding properties for adult hemoglobin. Molecular modeling studies which attempt to understand these properties of HbA2 are described.

KW - Allosteric regulator

KW - Delta gene

KW - Hemoglobin

KW - Tetramer strength

UR - http://www.scopus.com/inward/record.url?scp=0034466912&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034466912&partnerID=8YFLogxK

U2 - 10.1023/A:1007196118200

DO - 10.1023/A:1007196118200

M3 - Article

C2 - 11307949

AN - SCOPUS:0034466912

VL - 19

SP - 649

EP - 662

JO - Protein Journal

JF - Protein Journal

SN - 1572-3887

IS - 8

ER -