Expression, purification and characterization of Sorghum co-chaperone HOP ('Hsp70-Hsp90 Organizing Protein')

Author(s)

Ana Luíza Assin Squillace

• Document type: Master's Dissertation
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia
• Defense date: 2015-07-28

Examining board members

Carlos Henrique Inacio Ramos; Melissa Regina Fessel; Carlos Francisco Sampaio Bonafé

• Advisor: Carlos Henrique Inacio Ramos

Abstract

Molecular chaperones are a class of proteins that act by preventing aggregation and misfolding. In plants, the molecular chaperones have great importance because they have a role in protection against various types of environmental stresses. Since plants are sessile they need to respond efficiently to changes in temperature, salinity, drought, among others. The chaperone Hsp90 is essential for cell physiology, it is involved in the stabilization of various proteins, mostly involved in signaling. Its interaction with co-chaperones allows Hsp90 to act as a 'hub', a central point of regulation of several different proteins and regulatory pathways. Intervention in the interaction of Hsp90 co-chaperone is therefore a potentially effective strategy to modulate the function of this chaperone. One of its co-chaperones is Hop, that is present in many organisms, is able to associate directly to Hsp70 and Hsp90 mediating their association, which is dependent on ATP hydrolysis and exchange ADP / ATP. Hop consists of three domains with tetratricopeptides (TPR) repetions, involved in the interaction with chaperones. However, details about HOP oligomerization and mechanism of interaction are still missing in the literature. Recombinant Hop from sorgum (SbHop) was expressed in Escherichia coli, the protein was produced pure and was investigated by biophysical tools. Circular dichroism (CD) was used to investigate the secondary structure of the protein showing that it was produced folded and was predominantly ? helical. Thermal-induced unfolding experiments followed by CD and extrinsic fluorescence showed that the protein was stable up to 40°C. The monomeric state of Hop was showed by a technique combining gel filtration and light scattering. This study adds to a broad investigation on the mechanisms of plant stress tolerance (AU)

• FAPESP's process: 11/16047-0 - Study of the interaction between Hsp90 and co-chaperones with TPR domain.
• Grantee: Ana Luíza Assin Squillace
• Support Opportunities: Scholarships in Brazil - Master