A differential scanning calorimetric study of the bovine lens crystallins

Abstract

Differential scanning calorimetry was performed on the five major lens crystallin fractions [HM-alpha, alpha, beta H, beta L, and (beta s + gamma)] of the bovine lens as well as on more purified forms of alpha- and gamma-crystallins. All were found to be relatively thermally stable although the alpha-crystallin were found to at least partially unfold at an approximately 10 degrees C lower temperature than the beta and gamma fractions. Increasing protein concentration had little effect on gamma-crystallin thermograms but had marked effects on those of the alpha- and beta-crystallins. Increases in the thermal stability with increasing protein concentration for the beta-crystallins can be explained most simply by the known beta L/beta H equilibrium, but, in the case of the alpha-crystallins, excluded volume effects may be an important factor. In both cases, the increased stability at high concentrations could be of physiological relevance. As well as the expected endothermic unfolding transitions, all of the lens crystallins revealed exothermic peaks that correlate with protein precipitation. Interestingly, this phenomenon occurs only after extensive structural alteration in the case of the alpha-crystallins but is present very early in the initial stages of structural perturbation of the beta- and gamma-crystallins.

How can biochemical reactions within cells differ from those in test tubes?

“…Thus, the crystallin molecules that an animal is born with must remain structurally intact over much of its lifetime to preserve lens transparency. The thermal stability of these proteins increases with concentration (Steadman et al, 1989), and the extraordinary thermal stability of an intact lens has been attributed in part to the stabilizing effects of macromolecular crowding inside the lens cell (Bloemendal et al, 2004).…”

On the thermal stability of .alpha.-crystallin: z new insight from infrared spectroscopy

“…On the basis of the circular dichroism measurements of ellipticity at 217 nm as a function of temperature, it has been inferred that the protein retains its native secondary structure even at 100 "C (Maiti et al, 1988). Although the notion of a high thermal stability of a-crystallin has not been supported by a subsequent differential scanning calorimetry study of Steadman et al (1989), the latter work received surprisingly little attention. a-Crystallin is frequently referred to in the literature as the protein of extremely high thermodynamic stability.…”

Probing noncovalent structural features of proteins by mass spectrometry

“…Other methods for studying protein structure include fluorescence (74,75), phosphorescence (76,77), infrared (IR) (78-80), UV (81,82), and circular dichroism (CD) (83, 84) spectroscopies, and calorimetry (85,86). Although each of these methods has and continues to be used in studies of protein structure, none gives a detailed picture or structure, as is possible with X-ray diffraction or NMR spectroscopy.…”