Intrinsic tryptophan fluorescence spectra of wild type and mutant (R12A) Hsp27, αA- and αB-crystallin.

<p>Tryptophan fluorescence spectra of different samples (0.1 mg/ml protein) were recorded from 310–400 nm at 25°C. The excitation wavelength was 295 nm. Data were collected at 0.5 nm wavelength resolution.</p

Determination of the number of binding sites (n) and dissociation constant (K_d) values for the interaction of human Hsp27 and αA-and αB-crystallin and their R12A mutants with γ-crystallin at 60°C.

<p>Determination of the number of binding sites (n) and dissociation constant (K_d) values for the interaction of human Hsp27 and αA-and αB-crystallin and their R12A mutants with γ-crystallin at 60°C.</p

Identification of HI modification with the treatment of MGO detected by LC-MS/MS.

<p>Identification of HI modification with the treatment of MGO detected by LC-MS/MS.</p

Effect of R12A mutation on the chaperone function of Hsp27, αA- and αB-crystallin.

<p>The chaperone function of these three small heat shock proteins (wild type and mutants)was assessed using three client proteins, as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030257#s3" target="_blank"><i>Materials and Methods</i></a>. (<b>A</b>) Citrate synthase (CS); (<b>B</b>) γ-crystallin and (<b>C</b>) Lactate dehydrogenase (LDH).</p

Effect of R12A mutation on the surface hydrophobicity of Hsp27 and α-crystallin.

<p>The surface hydrophobicity of wild type and mutant proteins was estimated using a hydrophobic probe, TNS. Protein concentration was 0.1 mg/ml and TNS concentration was 100 µM. The fluorescence spectrum of TNS bound to different samples at 25°C was recorded from 350–520 nm. The excitation wavelength was 320 nm.</p

MGO reacts with proteins to form AGEs, like, hydroimidazolone, argpyrimidine and MOLD in tissue proteins.

<p>MGO reacts with proteins to form AGEs, like, hydroimidazolone, argpyrimidine and MOLD in tissue proteins.</p

Sequence alignment and SDS-PAGE of recombinant human Hsp27, αA- and αB-crystallin.

<p>(<b>A</b>) Amino-acid sequence alignment between these three small heat shock proteins was performed using the MULTIPLE SEQUENCE ALIGNMENT program (T-Coffee). (<b>B</b>) SDS-PAGE of purified proteins. M = Molecular weight markers.</p

Binding constant of wild type and R12A mutants of Hsp27, αA- and αB-crystallin for γ-crystallin.

<p>Binding parameters for the interaction between γ-crystallin and different small heat shock proteins at 60°C were estimated from Scatchard plot.</p

The C_1/2 and the ΔG⁰ values of the wild-type and R12A mutants of α-crystallin and Hsp27 at 25°C.

<p>The C_1/2 and the ΔG⁰ values of the wild-type and R12A mutants of α-crystallin and Hsp27 at 25°C.</p

The molar mass and the hydrodynamic radius of the wild-type and R12A mutants of α-crystallin and Hsp27.

<p>The molar mass and the hydrodynamic radius of the wild-type and R12A mutants of α-crystallin and Hsp27.</p