Metal-binding polymorphism in late embryogenesis abundant protein AtLEA4-5, an intrinsically disordered protein

Abstract

Late embryogenesis abundant (LEA) proteins accumulate in plants during adverse conditions and their main attributed function is to confer tolerance to stress. One of the deleterious effects of the adverse environment is the accumulation of metal ions to levels that generate reactive oxygen species, compromising the survival of cells. AtLEA4-5, a member of group 4 of LEAs in Arabidopsis, is an intrinsically disordered protein. It has been shown that their N-terminal region is able to undergo transitions to partially folded states and prevent the inactivation of enzymes. We have characterized metal ion binding to AtLEA4-5 by circular dichroism, electronic absorbance spectroscopy (UV–vis), electron paramagnetic resonance, dynamic light scattering, and isothermal titration calorimetry. The data shows that AtLEA4-5 contains a single binding site for Ni(II), while Zn(II) and Cu(II) have multiple binding sites and promote oligomerization. The Cu(II) interacts preferentially with histidine residues mostly located in the C-terminal region with moderate affinity and different coordination modes. These results and the lack of a stable secondary structure formation indicate that an ensemble of conformations remains accessible to the metal for binding, suggesting the formation of a fuzzy complex. Our results support the multifunctionality of LEA proteins and suggest that the C-terminal region of AtLEA4-5 could be responsible for antioxidant activity, scavenging metal ions under stress conditions while the N-terminal could function as a chaperone.

The in vitro structure and functions of the disordered late embryogenesis abundant three proteins

“…The upregulation of AtLEA3‐2 expression during oxidative stress suggests that metal binding may play a role in this protective mechanism 38 . First, the metal binding capabilities of the LEA3 proteins were analyzed since other LEA proteins have been shown to have this role 29,48 . The metal salts (CaCl 2 , MgCl 2 , CoCl 2 , FeCl 3 , ZnCl 2 , CuCl 2 , and NiCl 2 ) were chosen because of their physiological relevance to the mitochondrion, 49 and IMAC was used since it provides a rapid method to screen for metal binding.…”

Modular Assembly of Ordered Hydrophilic Proteins Improve Salinity Tolerance in Escherichia coli

“…Karamjeet et al predicted the conserved domains of plant G3LEA proteins and found that there are four N-terminal conserved motifs (called MAaRS, MARS, MGRX, and M [AS] [RK], respectively), which may be the signal for group 3 protein to locate mitochondria [ 39 ]. At LEA4-5, a member of group 4 of LEAs in Arabidopsis , it has been found that their N-terminal region can undergo a transition to a partially folded state and prevent enzyme inactivation, and, interestingly, the ability to gain structure under water limiting conditions is limited to the conserved region at the N-terminal, indicating the N-terminal could function as a chaperone [ 40 , 41 ]. The late embryogenesis abundant (LEA)-like protein CDeT11-24 of C. Plantagineum carries an N-terminal lysine-rich sequence, which is identified this region to be responsible for both activities: enzyme protection and phosphatidic acid interaction [ 42 ].…”

Investigation of a Novel Salt Stress-Responsive Pathway Mediated by Arabidopsis DEAD-Box RNA Helicase Gene AtRH17 Using RNA-Seq Analysis

“…LEA4-5 is a member of LEA proteins. The C-terminal region of LEA4-5 is responsible for its antioxidant activity and in the scavenging of metal ions under stress conditions, whereas the N-terminal can function as a chaperone in the folding of enzyme proteins and in preventing their unfolding that results in protection of the enzyme activity [19,30]. TSPO, belonging to the Trp-rich sensory protein/peripheral-type benzodiazepine receptor group protein, interacts with and regulates PIP2;7, a plasma membrane aquaporin, in the endoplasmic reticulum and Golgi membrane during abiotic stress conditions [20,21].…”