Three-dimensional structure of the truncated core of the Saccharomyces cerevisiae pyruvate dehydrogenase complex determined from negative stain and cryoelectron microscopy images
| Autor: | Xiao-Da Niu, John P. Schroeter, Steven J. Kolodziej, Timothy S. Baker, Lester J. Reed, James K. Stoops |
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| Rok vydání: | 1992 |
| Předmět: |
Biochemistry & Molecular Biology
Saccharomyces cerevisiae Proteins Macromolecular Substances 1.1 Normal biological development and functioning Pyruvate Dehydrogenase Complex Bioengineering Saccharomyces cerevisiae Biology Dihydrolipoyllysine-Residue Acetyltransferase Electron Medical and Health Sciences Biochemistry law.invention Structural Acetyltransferases Models Underpinning research law Freezing Microscopy Molecular Biology Staining and Labeling Resolution (electron density) Pyruvate Dehydrogenase (Lipoamide) Cell Biology Biological Sciences Pyruvate dehydrogenase complex Negative stain Recombinant Proteins Crystallography Chemical Sciences Electron microscope Macromolecule |
| Zdroj: | The Journal of biological chemistry, vol 267, iss 34 |
| ISSN: | 0021-9258 |
| DOI: | 10.1016/s0021-9258(18)35830-7 |
| Popis: | Dihydrolipoamide acyltransferase (E2), a catalytic and structural component of the three functional classes of multienzyme complexes that catalyze the oxidative decarboxylation of alpha-keto acids, forms the central core to which the other components are attached. We have imaged by negative stain and cryoelectron microscopy the truncated dihydrolipoamide acetyltransferase core (60 subunits; M(r) = 2.7 x 10(6)) of the Saccharomyces cerevisiae pyruvate dehydrogenase complex. Using icosahedral particle reconstruction techniques, we determined its structure to 25 A resolution. Although the model derived from the negative stain reconstruction was approximately 20% smaller than the model derived from the frozen-hydrated data, when corrected for the effects of the electron microscope contrast transfer functions, the reconstructions showed excellent correspondence. The pentagonal dodecahedron-shaped macromolecule has a maximum diameter, as measured along the 3-fold axis, of approximately 226 A (frozen-hydrated value), and 12 large openings (approximately 63 A in diameter) on the 5-fold axes that lead into a large solvent-accessible cavity (approximately 76-140 A diameter). The 20 vertices consist of cone-shaped trimers, each with a flattened base on the outside of the structure and an apex directed toward the center. The trimers are interconnected by 20 A thick "bridges" on the 2-fold axes. These studies also show that the highest resolution features apparent in the frozen-hydrated reconstruction are revealed in a filtered reconstruction of the stained molecule. |
| Databáze: | OpenAIRE |
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