Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.64 (MPP)
 1,876 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The aim was to examine the functional importance in the norepinephrine transporter (NET) of (i) the phenylalanine residue at position 531 in transmembrane domain (TMD) 11 by mutating it to tyrosine in the rat (rF531Y) and human (hF531Y) NETs and (ii) the highly conserved tyrosine residues at positions 249 in TMD 4 of human NET (hNET) (mutated to alanine: hY249A) and 271 in TMD 5, by mutating to alanine (hY271A), phenylalanine (hY271F) and histidine (hY271H). The effects of the mutations on NET function were examined by expressing the mutant and wildtype NETs in COS-7 cells and measuring the K(m) and V(max) for uptake of the substrates, [3H]norepinephrine, [3H]MPP(+) and [3H]dopamine, the K(D) and B(max) for [3H]nisoxetine binding and the K(i) of the inhibitors, nisoxetine, desipramine and cocaine, for inhibition of [3H]norepinephrine uptake. The K(m) values of the substrates were lower for the mutants at amino acid 271 than hNET and unaffected for the other mutants, and each mutant had a significantly lower V(max) than NET for substrate uptake. The mutations at position 271 caused an increase in the K(i) or K(D) values of nisoxetine, desipramine and cocaine, but there were no effects for the other mutations. Hence, the 271 tyrosine residue in TMD 5 is an important determinant of NET function, with the mutants showing an increase in the apparent affinities of substrates and a decrease in the apparent affinities of inhibitors, but the 249 tyrosine and 531 phenylalanine residues do not have a major role in determining NET function.
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PMID:Tyrosine residue 271 of the norepinephrine transporter is an important determinant of its pharmacology. 1174 60

Recently, another research group has reported an almost complete loss of function of the human norepinephrine transporter (hNET) in patients who had orthostatic intolerance and who were heterozygous for a guanine to cytosine exchange, resulting in a hNET Ala(457)Pro variant. To explore the reason for the deficiency in NET function, we compared in detail the pharmacology of the Ala(457)Pro variant with that of the wild-type hNET in COS-7 cells transiently transfected with hNET or Ala(457)Pro cDNA. Compared to the wild-type hNET, the Ala(457)Pro variant exhibited a five-fold higher affinity for cocaine, but a two-fold lower affinity for the NET inhibitor nisoxetine, and an unchanged affinity for the antidepressant desipramine. Plasma membrane expression (measured as Bmax of [3H]nisoxetine binding) of the Ala(457)Pro variant was only 40% of that of the wild-type hNET. The Ala(457)Pro variant showed a six- to 10-fold decrease in affinity for the substrates dopamine and 1-methyl-4-phenylpyridinium (MPP(+)). Compared with the wild-type hNET, the maximum rate (V(max)) of norepinephrine uptake by the Ala(457)Pro variant was slightly reduced, whereas the turnover number (calculated from V(max)/B(max)) was approximately two-fold higher. However, the Ala(457)Pro variant exhibited a 50-fold higher K(m) (i.e. lower apparent affinity) for norepinephrine than the wild-type hNET. Thus, the previously reported loss of function of the Ala(457)Pro variant associated with orthostatic intolerance is only partly due to a reduction in plasma membrane expression of the transporter, and is mainly caused by the pronounced reduction in the apparent affinity of norepinephrine.
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PMID:Pharmacological properties of the naturally occurring Ala(457)Pro variant of the human norepinephrine transporter. 1187 70