Recognition of protein substrates by protein-disulfide isomerase. A sequence of the b' domain responds to substrate binding
| Autor: | P Y, Cheung, J E, Churchich |
|---|---|
| Rok vydání: | 1999 |
| Předmět: |
Adenosine Triphosphatases
Protein Denaturation Protein Folding Protein Conformation Swine Myocardium Sulfhydryl Reagents Protein Disulfide-Isomerases Chaperonin 60 Fluoresceins Substrate Specificity Enzyme Activation Iodoacetamide Spectrometry Fluorescence Liver Malate Dehydrogenase Animals Trypsin Fluorescent Dyes Protein Binding |
| Zdroj: | The Journal of biological chemistry. 274(46) |
| ISSN: | 0021-9258 |
| Popis: | Refolding of partially folded mitochondrial malate dehydrogenase (mMDH) is assisted by protein-disulfide isomerase (PDI). The addition of a 20-fold molar excess of PDI over denatured protein (0. 1 microM) accelerates the recovery of catalytic activity. PDI fluorescence measurements show that 1 mol of PDI binds 1 mol of denatured mMDH when their concentrations approach 1 microM. The binding of PDI, derivatized with the fluorescence probe iodoacetamide fluorescein, to partially folded mMDH is characterized by a dissociation constant of 0.2 microM. It is shown that the fluorescence probe is covalently attached to a SH residue located in the b' domain. Based on the fluorescence measurements of native and derivatized PDI, it is suggested that recognition of the unfolded substrate involves conformational changes propagated to several domains of PDI. |
| Databáze: | OpenAIRE |
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