Studies of the N-terminal half of human lactoferrin produced from the cloned cDNA demonstrate that interlobe interactions modulate iron release.
J Biol Chem
; 267(20): 13857-62, 1992 Jul 15.
Article
em En
| MEDLINE
| ID: mdl-1378432
ABSTRACT
The factors influencing iron binding and release by lactoferrin have been addressed by comparison of the native full length molecule (Lf) with the N-terminal half of human lactoferrin (LfN) produced from the cloned cDNA expressed in baby hamster kidney (BHK) cells. The coding sequences for LfN were inserted into the expression vector pNUT between the metallothionein promoter and the human growth hormone transcription termination sequences. Transformed BHK cells were grown in roller bottles where concentrations of LfN as high as 35 mg/liter were obtained. The pure protein, produced by the transformed BHK cells, was characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, protein blotting and immunodetection, N-terminal sequence analysis, UV-visible spectroscopy, electron spin resonance spectroscopy, and measurements of metal binding and release. By these criteria LfN was found to be correctly processed, glycosylated, and able to bind iron reversibly. Both UV-visible and electron spin resonance spectra of the half molecule were very similar to those of native lactoferrin and the full length lactoferrin produced in BHK cells, but there were marked differences in the pH at which iron release occurred. Iron release from LfN occurs in the pH range 6.0-4.0, compared with 4.0-2.5 for native lactoferrin and 6.2-4.0 for transferrin. These results suggest that the more facile release of iron from LfN compared with native lactoferrin results from the absence of stabilizing contacts between the N- and C-terminal halves and that the characteristic difference in pH stability between lactoferrins and transferrins is due primarily to differences in these interactions.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
DNA
/
Ferro
/
Lactoferrina
Tipo de estudo:
Prognostic_studies
Limite:
Animals
/
Humans
Idioma:
En
Revista:
J Biol Chem
Ano de publicação:
1992
Tipo de documento:
Article
País de afiliação:
Nova Zelândia