The plasma membrane Ca2+ pump mutant lysine591 --> arginine retains some activity, but is still inactivated by fluorescein isothiocyanate.
Inactivation of the wild-type human plasma membrane Ca2+ pump (isoform 4b) by fluorescein isothiocyanate is accompanied by covalent modification of Lys591. The mutation of Lys591 to arginine reduced the Ca2+ transport activity to 35% of the wild-type, and diminished the amount of acylphosphate formed from ATP by a corresponding amount. When this mutant was treated with fluorescein isothiocyanate; the enzyme was still irreversibly inactivated, even though no reactive residue was available at position 591. The results show that, although Ca2+ pump function is sensitive to the residue at position 591, Lys591 is not essential for enzyme activity. They also demonstrate that irreversible inhibition of the plasma membrane Ca2+ pump by fluorescein isothiocyanate does not require the covalent modification of Lys591. This indicates that fluorescein isothiocyanate reacts with lysine residues at other positions in addition to Lys591.
Adamo
H P
HP
Instituto de Quimica y Fisicoquimica Biologica (UBA-CONICET), Facultad de Farmacia y Bioquimica, Buenos Aires, Argentina.
Filoteo
A G
AG
Penniston
J T
JT
D000255:Adenosine Triphosphate / Q000494:pharmacology; D001120:Arginine / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D016650:Fluorescein-5-isothiocyanate / Q000494:pharmacology*; D006801:Humans; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010766:Phosphorylation
D000255:Adenosine Triphosphate; D001120:Arginine; D000252:Calcium-Transporting ATPases; D016650:Fluorescein-5-isothiocyanate; D008239:Lysine