Effect of ions and lipids on ATPase activity of the Spf1 P5- ATPase from Saccharomyces cerevisiae

PETROVICH GUIDO DANIEL; CORRADI GERARDO RAUL; ADAMO HUGO PEDRO

La Plata

Congreso; XLVII Reunión Anual de la Sociedad Argentina de BiofísicaXLVII Reunión Anual de la Sociedad Argentina de Biofísica; 2018

Sociedad Argentina de Biofísica

P-type  ATPases (P1 to P5) are integralmembraneproteins that  transport differentligands againstits concentration gradient,through ATP hydrolysis. P5-ATPases arethe  least  studied membersof  the  whole  family of  P-ATPases and its transported substrate   remains unknown.  Several    human P5-ATPases  are  implicated inneurological disorders,  as  the   Kufor-Rakeb  syndrome,   a   parkinsonism with dementia, hereditaryspastic paraplegia, neuronal ceroid lipofuscinosis, autism and intellectual disability. With the aim of advancingthe  knowledge of P5-ATPases wehave   investigated the effect of  lipids  and ions  on  the  ATPase  activity of  therecombinant Spf1 P5-ATPase of Saccharomyces cerevisiae.The  highest levels  ofATPase  activity were  achieved   when  asolectin  from   soybean  was   used   to supplement the micelar preparationof the  protein indicating that  asolectin makesthe best  micelar environmentfor  Spf1  ATP hydrolysis .  While monovalent  ions didn?t  affect  ATPase  activity Spf1,   it was   inhibited by mercury,  aluminum,lanthanum  and   cadmium. Like   other  P-ATPases, Spf1  required magnesium to hydrolyze ATP. The dependence of Spf1 ATPase activity with  Mg 2+ was best fittedby a double hyperbola, afact  that could  indicate the existence of two binding sites for  magnesium. On the other hand,  high  concentration of other divalentcations reduced  the   ATPase  activity. Interestingly,  the  Spf1  ATPase  had  a  biphasicdependency with  zinc. At low concentrations zinc increased the ATPase while  itproduced inhibition at  higher concentrations. High  inhibitory concentrations zincin the presence ofATP changedthe sensibility of Spf1 to chymotrypsin suggesting the stabilization of an inhibited conformation.