KINETIC CHARACTERIZATION OF THE RECONSTITUTED CARNITINE CARRIER FROM RAT-LIVER MITOCHONDRIA

被引:65
作者
INDIVERI, C
TONAZZI, A
PREZIOSO, G
PALMIERI, F
机构
[1] UNIV BARI,DEPT PHARMACOBIOL,BIOCHEM LAB,200 RE DAVID 4,I-70125 BARI,ITALY
[2] CNR,STUDY MITOCHONDRIA & BIOENERGET UNIT,I-70126 BARI,ITALY
关键词
CARNITINE CARRIER; RECONSTITUTION; KINETICS; LIPOSOME; MITOCHONDRION; (RAT LIVER);
D O I
10.1016/0005-2736(91)90235-Z
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The carnitine carrier was purified from rat liver mitochondria and reconstituted into liposomes by removing the detergent from mixed micelles by Amberlite. Optimal transport activity was obtained with 1-mu-g/ml and 12.5 mg/ml of protein and phospholipid concentration, respectively, with a Triton X-100/phospholipid ratio of 1.8 and with 16 passages through the same Amberlite column. The activity of the carrier was influenced by the phospholipid composition of the liposomes, being increased in the presence of cardiolipin and decreased in the presence of phosphatidylinositol. In the reconstituted system the incorporated carnitine carrier catalyzed a carnitine/carnitine exchange which followed a first-order reaction. The maximum transport rate of external [H-3]carnitine was 1.7 mmol/min per g protein at 25-degrees-C and was independent of the type of countersubstrate. The half-saturation constant (K(m)) for carnitine was 0.51 mM. The affinity of the carrier for acylcarnitines was in the mu-M range and depended on the carbon chain length. The activation energy of the carnitine/carnitine exchange was 133 kJ/mol. The carrier function was independent of the pH in the range between 6 and 8 and was inhibited at pH below 6.
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页码:231 / 238
页数:8
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