Effects of heme addition on formation of stable human globin chains and hemoglobin subunit assembly in a cell-free system.
Our previous assembly studies to form hemoglobin hetero-dimers and -tetramers using a coupled transcription/translation cell-free system suggested that alpha-globin chains bind to nascent non-alpha chains during and/or soon after translation to promote hemoglobin formation [Adachi et al., J. Biol Chem. 2002 (277) 13415]. In this report effects of CN-hemin on subunit assembly were studied using this cell-free system. Addition of CN-hemin and excess unlabeled heme-containing partner chains during synthesis leads to formation of radiolabeled heme-containing alpha(h)beta(h) hetero-dimers. In contrast, in the absence of added CN-hemin, unlabeled heme-containing alpha or beta chains can assemble with newly synthesized radiolabeled beta- and alpha-globin chains to form heme-containing alpha(h)beta(h) and semi-alpha (alpha(h)beta(0)) or semi-beta (alpha(0)beta(h)) hetero-dimers, respectively. These results suggest the existence of semi-hemoglobins as intermediates prior to formation of heme-containing alpha(h)beta(h) and indicate transfer of heme from alpha and/or beta chains into semi-hemoglobin hetero-dimers to form heme-containing hetero-dimers.
Adachi
Kazuhiko
K
The Children's Hospital of Philadelphia, Division of Hematology and University of Pennsylvania School of Medicine, 19104, USA. adachi@email.chop.edu
Zhao
Yi
Y
Surrey
Saul
S
D002474:Cell-Free System; D018076:DNA, Complementary / Q000235:genetics; D019281:Dimerization; D005822:Genetic Vectors; D005914:Globins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006418:Heme / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006454:Hemoglobins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D066298:In Vitro Techniques; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D018076:DNA, Complementary; D006454:Hemoglobins; D021122:Protein Subunits; D011994:Recombinant Proteins; D006418:Heme; D005914:Globins