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J Horwitz

Publications and source records attributed to J Horwitz.

204 records · Page 12Linked to original sources
PubMedDate not supplied

Mutation of alpha B-crystallin: effects on chaperone-like activity.

A recent paper by Plater et al. [20], showed that the mutation of a single phenylalanine residue F27R in mouse alpha B completely abolished the chaperone-like property of alpha-crystallin when assayed with insulin at 25 degrees C or with gamma-crystallin at 66 degrees C. We have produced the same mutation as well as some additional mutations in human alpha B-crystallin. Our data suggest that the F27R mutation effected the thermal stability of alpha B-crystallin making it unstable at temperatures > or = 60 degrees C. In agreement with the published work, at these temperatures the F27R human recombinant alpha B-crystallin does not protect the target protein from aggregation. When assayed with insulin or alpha-lactalbumin at 25 or 37 degrees C, however, there were no differences in the protective abilities between the native alpha B-crystallin or the F27R mutated human alpha B-crystallin. Several other multiple mutations involving proline residues were also produced. These mutations did not effect the chaperone-like properties of human alpha B-crystallin, but some of them did effect the native molecular weight size as judged by gel filtration chromatography.

Animals↗
PubMedDate not supplied

Biochemical analysis of microdissected sections from the normal and cataractous human lens.

To better compare possible changes in soluble protein content during human cataractogenesis, microdissected sections from opaque and highly pigmented versus transparent parts of the lens were analyzed by high pressure liquid chromatography, isoelectric focusing, sodium dodecylsulfate polyacrylamide gel electrophoresis, and tryptic mapping of radioiodinated peptides. Using this methodology, resolution of proteins and their tryptic peptides can be obtained from individual lens sections containing as little as 5-10 micrograms of protein. Furthermore, the origin of each protein analyzed by these methods can be verified by comparison with tryptic peptide maps of previously purified human crystallins. Included are examples of how this technique can be used to characterize differences in protein composition from opaque versus transparent sections from different human lenses, as well as brunescent vs lightly-pigmented sections from the same human lens.

Autoradiography↗
PubMedDate not supplied

Comparison of in vitro proteolysis products of the normal human lens with putative in vivo proteolysis products of the cataractous lens.

Treatment of normal lens homogenates with trypsin results in the production of polypeptides of approximately 4,000 and 8,000 daltons. Peptide maps of these components are very similar to poly-peptides of comparable molecular weight found only in cataracts with brunescent and opaque nuclei. Together, these results demonstrate that putative proteolysis products produced in vivo during human cataractogenesis can be mimiced in vitro by the addition of exogeneous proteases to the normal lens homogenate.

Cataract↗