Real-time heterogeneous protein-protein interaction between αA-crystallin N-terminal mutants and αB-crystallin using quartz crystal microbalance (QCM).

Abstract:

:The lens transparency depends on higher concentration of lens proteins and their interactions. α-Crystallin is one of the predominant lens proteins, responsible for proper structural and functional architecture of the lens microenvironment, and any alteration of which results in cataract formation. The R12C, R21L, R49C and R54C are the most significant and prevalent αA-crystallin congenital cataract-causing mutants worldwide. Protein-protein interaction, crucial for lens proper structure and function, was posited to be lost due to point mutation and the elucidation of which could shed light on the molecular basis of cataract. In this conjuncture, we report quartz crystal microbalance (QCM) as a warranted technique for real-time analysis of protein-protein interaction between the N-terminal mutants of αA-crystallin and αB-crystallin. The biophysical characteristics of the mutated proteins were determined by size-exclusion HPLC, far-UV circular dichroism and fluorescence studies. Far-UV circular dichroism spectral analysis displayed slight modifications in β-sheet of R54C mutant. Altered intrinsic tryptophan fluorescence and decreased bis-ANS fluorescence were observed in all the N-terminal mutations revealing the tertiary structural changes and decreased exposure of surface hydrophobicity. An emphatic fall in the chaperone activity was observed in the N-terminal mutants, R12C, R21L and R54C. QCM analysis revealed the occurrence of strong heterogeneous interaction between αA-crystallin and αB-crystallin. Nevertheless, decreased interactions were observed with the N-terminal mutants. In summary, the present study concludes that the loss of interactions between αA-crystallin N-terminal mutants and αB-crystallin signifies quaternary structural alterations due to mutation in the arginine residues.

journal_name

Amino Acids

journal_title

Amino acids

authors

Ramkumar S,Fujii N,Sakaue H,Fujii N,Thankappan B,kumari RP,Natarajaseenivasan K,Anbarasu K

doi

10.1007/s00726-015-1935-z

subject

Has Abstract

pub_date

2015-05-01 00:00:00

pages

1035-43

issue

5

eissn

0939-4451

issn

1438-2199

journal_volume

47

pub_type

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