Determination of nicotinamide phosphoribosyltransferase activity in human erythrocytes: high-performance liquid chromatography-linked method.
A radiochemical reverse-phase-high-performance liquid chromatography-linked method to measure the activity of nicotinamide phosphoribosyltransferase (EC 2.4.2.12) in crude lysates of human red blood cells is described. The apparent Km for nicotinamide was in the micromolar range, much lower than that described in human erythrocytes in the past. The enzyme activity in crude hemolysates was found to be extremely low (21 +/- 3.5 nmol x h-1 x g-1 Hb); nevertheless, the low Km for nicotinamide might account for the production of pyridine nucleotides reported by us for intact erythrocytes incubated at low, physiological concentrations of this substrate.
Rocchigiani
M
M
Dipartimento di Biologia Molecolare, Università di Siena, Italia.
Micheli
V
V
Duley
J A
JA
Simmonds
H A
HA
D002250:Carbon Radioisotopes; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D054409:Nicotinamide Phosphoribosyltransferase; D010430:Pentosyltransferases / Q000097:blood*
D002250:Carbon Radioisotopes; D010430:Pentosyltransferases; D054409:Nicotinamide Phosphoribosyltransferase