当前位置： SCI文献检索 > BIOCHEMISTRY期刊下所有文献 > Structural and functional properties of human hemoglobins reassembled after synthesis in Escherichia coli.

Structural and functional properties of human hemoglobins reassembled after synthesis in Escherichia coli.

Abstract：

:Human hemoglobin produced in the Escherichia coli coexpression system of Hernan et al. [(1992) Biochemistry 31, 8619-8628] has been transformed into a functionally homogeneous protein whose properties closely approximate those of normal hemoglobin A. Both of the alpha and beta chains of this hemoglobin contain a valine-methionine substitution at position 1 in order to accommodate the difference in specificity of the protein-processing enzymes of procaryotes. Despite extensive purification, functional homogeneity of the E. coli expressed hemoglobin was achieved only by the complete disassembly of the hemoglobin into its component alpha and beta globins and their reassembly in the presence of hemin. The kinetics of CO combination and the thermodynamics of O2 binding and cooperativity of the reassembled alphaV1M-betaV1M hemoglobin closely approximate those of HbA. The alpha globin obtained from the E. coli expressed hemoglobin was also combined with normal human beta chains and hemin to form the alphaV1M variant. The alpha+M variant of HbA, in which the normal N-terminal valine of the alpha chains is preceded by a methionine residue, was prepared by the same procedure. The kinetics of the reactions of CO with the alphaV1M and alpha+M variants are similar to those for HbA. The equilibria of oxygen binding to alphaV1M and HbA are similar whereas alpha+M exhibits a significantly higher oxygen affinity. The three-dimensional structures of alphaV1M and alpha+M offer an explanation for the latter affinity difference. Although the structures of alphaV1M and HbA, which have been determined by X-ray crystallography, are virtually indistinguishable except at the N-terminal residues, that of alpha+M indicates the displacement of a solvent molecule, possibly a chloride ion, from arginine 141alpha. Such an alteration in an anion binding site could result in increased oxygen affinity.

journal_name

Biochemistry

journal_title

Biochemistry

authors

Hui HL,Kavanaugh JS,Doyle ML,Wierzba A,Rogers PH,Arnone A,Holt JM,Ackers GK,Noble RW

doi

10.1021/bi981986g

subject

Has Abstract

pub_date

1999-01-19 00:00:00

pages

1040-9

issue

eissn

0006-2960

issn

1520-4995

pii

bi981986g

journal_volume

pub_type

杂志文章

在线工具

Structural and functional properties of human hemoglobins reassembled after synthesis in Escherichia coli.