Addition of αA-Crystallin Sequence 164–173 to a Mini-Chaperone DFVIFLDVKHFSPEDLT Alters the Conformation but Not the Chaperone-like Activity

نویسندگان

  • Murugesan Raju
  • Puttur Santhoshkumar
  • Leike Xie
  • K. Krishna Sharma
چکیده

It has been shown that αA-mini-chaperone, a peptide representing the chaperone binding site in αA-crystallin, prevents destabilized protein aggregation. αA-Mini-chaperone has been shown to form amyloid fibrils. This study was undertaken to improve the stability of αA-mini-chaperone while preserving its anti-aggregation activity by fusing the flexible and solvent-exposed C-terminal 164-173 region of αA-crystallin to the mini-chaperone sequence DFVIFLDVKHFSPEDLT. The resulting chimeric chaperone peptide, DFVIFLDVKHFSPEDLTEEKPTSAPSS (designated CP1), was characterized. Circular dichroism studies showed that unlike αA-mini-chaperone with its β-sheet structure, the CP1 peptide exhibited a random structure. Transmission electron microscopy (TEM) examination of the CP1 peptide incubated in a shaker at 37 °C for 72 h did not reveal amyloid fibrils, whereas αA-mini-chaperone showed distinct fibrils. Consistent with TEM observation, the thioflavin T binding assay showed an increased level of dye binding in the mini-chaperone incubated at 37 °C and subjected to shaking but not of the CP1 peptide incubated under similar conditions. The chaperone activity of the CP1 peptide was comparable to that of αA-mini-chaperone against denaturing alcohol dehydrogenase, citrate synthase, and α-lactalbumin. Transduction of both peptide chaperones to COS-7 cells showed no cytotoxic effects. The antioxidation assay involving the H2O2 treatment of COS-7 cells revealed that αA-mini-chaperone and the CP1 peptide have comparable cytoprotective properties against H2O2-induced oxidative damage in COS-7 cells. This study therefore shows that the addition of C-terminal sequence 164-173 of αA-crystallin to αA-mini-chaperone influences the conformation of αA-mini-chaperone without affecting its chaperone function or cytoprotective activity.

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Correction to Addition of αA-Crystallin Sequence 164–173 to a Mini-Chaperone DFVIFLDVKHFSPEDLT Alters the Conformation but Not the Chaperone-like Activity

Received: May 12, 2014 Published: May 22, 2014 Figure 5. Aggregations of denaturing proteins in the presence of miniαA-chaperone or chimeric mini-chaperone (CP1). (A) Heatand EDTA-induced ADH (5 μM) aggregation in the presence of mini-αA (40 μM) or mini-αA (Δ87−88, 40 μM) at 37 °C. (B) Heat-induced citrate synthase (4 μM) aggregation assay at 43 °C in the presence of mini-αA (40 μM) or CP1 (40 ...

متن کامل

αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin

BACKGROUND A substitution mutation in human αA-crystallin (αAG98R) is associated with autosomal dominant cataract. The recombinant mutant αAG98R protein exhibits altered structure, substrate-dependent chaperone activity, impaired oligomer stability and aggregation on prolonged incubation at 37 °C. Our previous studies have shown that αA-crystallin-derived mini-chaperone (DFVIFLDVKHFSPEDLTVK) fu...

متن کامل

Molecular Mechanism of the Chaperone Function of Mini-α-Crystallin, a 19-Residue Peptide of Human α-Crystallin

α-Crystallin is the archetypical chaperone of the small heat-shock protein family, all members of which contain the so-called "α-crystallin domain" (ACD). This domain and the N- and C-terminal extensions are considered the main functional units in its chaperone function. Previous studies have shown that a 19-residue fragment of the ACD of human αA-crystallin called mini-αA-crystallin (MAC) show...

متن کامل

Chaperone peptides of α-crystallin inhibit epithelial cell apoptosis, protein insolubilization, and opacification in experimental cataracts.

α-Crystallin is a member of the small heat-shock protein (sHSP) family and consists of two subunits, αA and αB. Both αA- and αB-crystallin act as chaperones and anti-apoptotic proteins. Previous studies have identified the peptide (70)KFVIFLDVKHFSPEDLTVK(88) in αA-crystallin and the peptide (73)DRFSVNLDVKHFSPEELKVK(92) in αB-crystallin as mini-chaperones. In the human lens, lysine 70 (Lys(70)) ...

متن کامل

Role of Molecular Interactions and Oligomerization in Chaperone Activity of Recombinant Acr from Mycobacterium tuberculosis

Background: The chaperone activity of Mycobacterium tuberculosis Acr is an important function that helps to prevent misfolding of protein substrates inside the host, especially in conditions of hypoxia. Objectives: The aim of this study was to establish the correlation of structure and function of recombinant Acr proteins both before and after ge...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

عنوان ژورنال:

دوره 53  شماره 

صفحات  -

تاریخ انتشار 2014