Assembly of human hemoglobin (Hb) beta- and gamma-globin chains expressed in a cell-free system with alpha-globin chains to form Hb A and Hb F.
Rates of in vitro synthesis of radiolabeled gamma and beta chains made in a cell-free transcription/translation system were similar, but expressed globin chains were unstable. The addition of unlabeled beta or gamma chains at the start of chain synthesis generated radiolabeled beta(4) or gamma(2) and gamma(4) chains, respectively. If unlabeled alpha-globin chains were added at the start of chain synthesis, then approximately equal amounts of radiolabeled alphabeta or alphagamma bands were generated. If unlabeled Hb A or Hb F was added to reactions containing radiolabeled alphabeta or alphagamma prior to electrophoresis, then radiolabeled Hb A or Hb F tetramers, respectively, were generated. If alpha chains were added after synthesis of radiolabeled gamma chains made in the presence of unlabeled gamma chains, then little radiolabeled alphagamma formed. In contrast, if alpha chains were added after synthesis of radiolabeled beta chains made in the presence of unlabeled beta chains, then radiolabeled alpha(2)beta(2) formed. These findings suggest that beta and gamma chains associate with alpha chains during or soon after translation. This would prevent the formation of unstable monomers as well as stable gamma(2) dimers and suggests that alpha chains may bind to nascent non-alpha chains, acting as folding catalysts to promote functional tetrameric hemoglobin formation in vivo.
Adachi
Kazuhiko
K
Children's Hospital of Philadelphia, Division of Hematology and the University of Pennsylvania School of Medicine Philadelphia, Pennsylvania 19104, USA. adachi@email.chop.edu
Zhao
Yi
Y
Surrey
Saul
S
D002474:Cell-Free System; D018076:DNA, Complementary / Q000378:metabolism; D019281:Dimerization; D005319:Fetal Hemoglobin / Q000737:chemistry*; D005822:Genetic Vectors; D006441:Hemoglobin A / Q000737:chemistry*; D006454:Hemoglobins / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D008956:Models, Chemical; D011485:Protein Binding; D014176:Protein Biosynthesis; D013997:Time Factors; D014908:Triticum / Q000378:metabolism
D018076:DNA, Complementary; D006454:Hemoglobins; D006441:Hemoglobin A; D005319:Fetal Hemoglobin