Determination of ferric heme-human serum albumin by 1H NMR relaxometry.
A new method for the accurate determination of ferric heme-human serum albumin (heme-HSA) at concentrations down to the physiological level, i.e., in the micromolar concentration range, is proposed. This method is based on the (1)H NMR relaxometric properties of heme-HSA. Actually, the binding of the paramagnetic ferric heme to the primary binding site of HSA determines a strong paramagnetic enhancement of the water (1)H NMR relaxation rate. Although a linear relationship may be seen by operating at 20 MHz on conventional electromagnets, the method here reported is improved by working at 0.02 MHz on a field-cycling instrument. This (1)H NMR relaxometric method does not suffer from the presence in serum of heme catabolites (e.g., bilirubin) that affect significantly the optical determination of ferric heme-HSA in the micromolar concentration range. Paramagnetic ferric hemoglobin contribution may be selectively quenched by cyanide binding.
Fasano
Mauro
M
Dipartimento di Biologia Strutturale e Funzionale, Università dell'Insubria, Varese, Italy. mauro.fasano@uninsubria.it
Baroni
Simona
S
Aime
Silvio
S
Mattu
Marco
M
Ascenzi
Paolo
P
D006418:Heme / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D012709:Serum Albumin / Q000737:chemistry* / Q000378:metabolism
D012709:Serum Albumin; D006418:Heme