Conformational constraints for protein self-cleavage in the proteasome.
The proteasome is the central enzyme of protein degradation in the cytosol and the nucleus. It is involved in the removal of abnormal, misfolded or incorrectly assembled proteins, in the processing or degradation of transcriptional regulators in stress response, in degradation of cyclins in cell-cycle control, in the destruction of transcription factors or metabolic enzymes in cell differentiation and metabolic response, and in MHC class I mediated cellular immune response. By the analysis of the crystal and molecular structures of the 20 S proteasomes from the archaeon Thermoplasma acidophilum and from yeast it was shown that the beta-type subunits in which the proteolytic activities reside are members of the N-terminal nucleophile (Ntn) protein family. They are synthesized as proproteins and become active by autoprocessing at a Gly-1-Thr1 bond. The Thr1Ala mutant of subunit beta1/Pre3 of the 20 S proteasome from yeast is unable to autolyse. Its crystal and molecular structure at 2.2 A resolution described here shows that the pro-segment adopts a well-defined gamma-turn conformation at Gly-1 and provides a first view at an autolysis site in Ntn hydrolases. The Gly-1 carbonyl oxygen displays two hydrogen bonds. The modelled Thr1 side-chain is located above the gamma-turn bulge such that addition of its nucleophilic hydroxyl group to the electrophilic Gly-1 carbonyl carbon atom may proceed by very small motions. The pro-segment binding site and the catalytic site provide a rigid structural framework and appropriate hydrogen bond donors for this reaction. The same structure also supports addition of the Thr1 hydroxyl group to the carbonyl carbon atom of Leu-2 as a model for the first step in substrate hydrolysis by the proteasome.
Ditzel
L
L
Max-Planck-Institut für Biochemie, Martinsried, D-82152, Germany.
Huber
R
R
Mann
K
K
Heinemeyer
W
W
Wolf
D H
DH
Groll
M
M
D019843:Archaeal Proteins / Q000737:chemistry*; D018360:Crystallography, X-Ray; D003546:Cysteine Endopeptidases / Q000737:chemistry*; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry*; D046988:Proteasome Endopeptidase Complex; D011487:Protein Conformation; D011506:Proteins / Q000378:metabolism; D013822:Thermoplasma / Q000737:chemistry; D015003:Yeasts
D019843:Archaeal Proteins; D009097:Multienzyme Complexes; D011506:Proteins; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex