Characteristics of a Ca2+-ATPase activity in islet homogenates

59. ROSSI J.P.F.C., GRONDA, C.M. FERNÁNDEZ, H.N., Y GAGLIARDINO, J.J.

BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES

ELSEVIER SCIENCE BV

Lugar: Amsterdam; Año: 1988 vol. 943 p. 175 - 182

0005-2736

Ca2+-ATPase act~vity was measured in rat islet ltomogenate~ in a mediul of low ionic slrenSth cottlaillillg alow ¢o~entralimt 04[ Ca z+ and Mg 2+ ~ devoid of K +. The enzyme aclivily was highly selzsitlve toinhibition by ¢,omflpotma 48/80 (a calmedulbt inhibitor), stimulated by 120 ~ calm0sa~ln and slightlyaifeeted ~ i0 mM Na_,~, The addition M Mg ~÷ to the assay medlum promotes the ~aSal~manee Mapparent CaZ÷-ATPase aetivi~. Ouabain (0.t raM) did mt modify this ATPase aeflvlty. The enzyme sZmwedtwo kinetic ¢omlaments for Ca z+ as well as fro" ATP: one with high at~a'ent i~iinity aml loftvelocity and the other with low skqmrent affinity and high maximum velocity. Inmbalion el Islet Immogenatesin this assay medium with I¥-~PIATP in the px'sence of Im~eelytic tnldbitecs, results in theappearance of a s~agle labelled band of 130 kDa, identllled by gel eleetroplmresis. The incorporation of ~2pinto this band was similar in the la'esence of either 2.8 or 50 #tM Ca z+ and suscel~'ble to hydroxylamlneattack. The results indicate that, under the eoadl6em deseatbed above, the Ca2÷-ATPase. aetivity evtdenc~in the Islet homogeaales had eharaeteristks tesembllng those of the em.yme which ,'~J~lyzes the ~4atwardCa 2÷ trenslmrt. On the other hand, the melhod co~d provide a useful tool to test the elfect of differentagents which Mfe~ insulin seeretioB upon the islet plasma mem~ Ca~+-ATPase activity