Methods to investigate complex protein mixtures have been improved: Electrophoresis and staining of proteins from cerebrospinal fluid (CSF) and brain tissue have increased detection from 300 to 1500 and 1000 to 3500 respectively. Disease-associated proteins in the CSF have been further investigated. Purification and partial amino acid sequences have been obtained from 6 proteins: One of the proteins is identified as a transthyretin molecule, twice the molecular weight of the usual protein, and absent form the plasma. The other 5 proteins are not homologous to any of the known sequences in the NBRF protein database. Of these 5, two are diagnostically important in Creutzfeldt-Jakob disease, one is reduced in schizophrenia, and tow are increased in multiple sclerosis. Synthetic peptides have been made to these peptides, and antibodies are being produced, in order to develop sensitive rapid and accurate immunoassays for these newly characterized proteins. Two-dimensional electrophoretic survey studies have been continued with various disease model systems. Brains from inbred m ice strains have been studied, with a normal protein reference being established for common strains, in order to study neurological mutants that have been bred on a similar genetic background. Hamster brain proteins have been studied, in norma, heterozygotic autosomal dominant circadian rhythm mutants and homozygotic mutants. Multiple protein alterations have been identified, and further investigations are in progress to define which of these proteins is associated with variations in circadian rhythm. Further developments to advance the resolution and detection of proteins has resulted in the synthesis of a new crosslinker which allows the polymerization of gels with better physical strength, less hydrophilicity, and better protein separation by two dimensional gel electrophoresis. Its probable lower affinity for silver ions and its greater resistance to hydrolysis also delays the appearance of a background stain with ammoniacal silver nitrate.
| Zullo, S J; Parks, W T; Chloupkova, M et al. (2005) Stable transformation of CHO Cells and human NARP cybrids confers oligomycin resistance (oli(r)) following transfer of a mitochondrial DNA-encoded oli(r) ATPase6 gene to the nuclear genome: a model system for mtDNA gene therapy. Rejuvenation Res 8:18-28 |
