Structural Changes beyond the EF-Hand Contribute to Apparent Calcium Binding Affinities: Insights from Parvalbumins

2021 ◽  
Author(s):  
Kalyan Immadisetty ◽  
Bin Sun ◽  
Peter M. Kekenes-Huskey
Keyword(s):  
Calcium Binding ◽  
Structural Changes ◽  
Binding Affinities ◽  
Ef Hand

scholarly journals Machine learning-based modulation of Ca2+-binding affinity in EFhand proteins and comparative structural insights into site-specific cooperative binding

2021 ◽  
Author(s):  
Mohit Mazumder ◽  
Sanjeev Kumar ◽  
Devbrat Kumar ◽  
Samudrala Gourinath
Keyword(s):  
Calcium Binding ◽  
Site Directed Mutagenesis ◽  
Binding Affinities ◽  
Dynamic Correlation ◽  
Loop Sequence ◽  
Binding Isotherms ◽  
Ef Hand ◽  
Coordination Spheres ◽  
Living Organisms ◽  
Almost All

Ca2+-binding proteins are present in almost all living organisms and different types display different levels of binding affinities for the cation. Here, we applied two new scoring schemes enabling the user to manipulate the binding affinities of such proteins. We specifically designed a unique EF-hand loop capable of binding calcium with high affinity by altering five residues of the loop based on the scoring scheme. We worked on the N-terminal domain of Entamoeba histolytica calcium-binding protein1 (NtEhCaBP1), and used site-directed mutagenesis to incorporate the designed loop sequence into the second EF hand motif of this protein. The binding isotherms calculated using ITC calorimetry showed a ~500-fold greater association constant (Ka) for the mutant. The crystal structure of the mutant was also determined, and displayed more compact Ca2+-coordination spheres in both of its EF loops than did the structure of the wildtype protein, consistent with the greater calcium-binding affinities of the mutant. The NtEhCaBP1 mutant was also shown to form a hexamer rather than just a trimer, and this hexamer formation was attributed to the position of the last helix of the mutant having been changed as a result of the strong calcium coordination. Further dynamic correlation analysis revealed that the mutation in the second EF loop changed the entire residue network of the monomer, resulting in a stronger coordination of Ca2+.

Obtaining Site-Specific Calcium-Binding Affinities Of Calmodulin

2003 ◽  
Vol 10 (4) ◽  
pp. 331-345 ◽  
Author(s):  
Jenny Yang ◽  
Amy Gawthrop ◽  
Yiming Ye
Keyword(s):  
Calcium Binding ◽  
Binding Affinities ◽  
Site Specific
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