THE ATPASE ACTIVITY AND ASSEMBLY OF CLPB3/HSP100 FROM ARABIDOPSIS THALIANA CHLOROPLASTS

PARCERISA, IVANA L.; ROSANO, GERMÁN L.; CECCARELLI, EDUARDO A.

Mar del Plata

Congreso; LI Reunión Anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2015

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

To counteract protein aggregation, bacteria, fungi and plants contain a bi-chaperone system composed of ATPdependent Hsp70 and ClpB/Hsp100 chaperones, which rescue aggregated proteins and provide thermotolerance tocells. The ClpB component of this system is an AAA+ ATPase that forms an hexameric functional ring-like structureof identical protomers, each one containing two nucleotide binding domains (NBD-1 and -2). ClpB oligomerize inresponse to different stimuli such as nucleotide binding or ionic strength, which in turn regulate the activity of thechaperone. In this work, we provide the first characterization of the ClpB3/Hsp100 from A. thaliana chloroplasts.Oligomerization in the presence of ATP was studied by SEC and the innate ATPase activity analyzed under severalconditions by the Malachite Green method. We have established that ATP stimulates hexamerization while high ionicstrength favors disassembly of the complexes. Furthermore, we have determined the kinetic parameters of the enzymeand its activity under a range of pH values and temperatures. ClpB3 lacks activity at pH values equal to or lower than4.5 and has an optimal activity at temperatures around 62oC. Since the overall behavior of the enzyme resembles thatof bacterial and yeast homolog, these results suggest that ClpB3 from A. thaliana could act similarly in theacquisition of plant thermotolerance