Kinetic evidences for facilitation of peptide channelling by the proteasome activator PA28
Open Access
- 1 October 2000
- journal article
- research article
- Published by Wiley in European Journal of Biochemistry
- Vol. 267 (20), 6221-6230
- https://doi.org/10.1046/j.1432-1327.2000.01706.x
Abstract
The activation kinetics of constitutive and IFNγ‐stimulated 20S proteasomes obtained with homomeric (recPA28α, recPA28β) and heteromeric (recPA28αβ) forms of recombinant 11S regulator PA28 was analysed by means of kinetic modelling. The activation curves obtained with increasing concentrations of the individual PA28 subunits (RecP28α/RecP28β/RecP28α+ RecP28β) exhibit biphasic characteristics which can be attributed to a low‐level activation by PA28 monomers and full proteasome activation by assembled activator complexes. The dissociation constants do not reveal significant differences between the constitutive and the immunoproteasome. Intriguingly, the affinity of the proteasome towards the recPA28αβ complex is about two orders of magnitude higher than towards the homomeric PA28α and PA28β complexes. Striking similarities can been revealed in the way how PA28 mediates the kinetics of latent proteasomes with respect to three different fluorogenic peptides probing the chymotrypsin‐like, trypsin‐like and peptidylglutamyl‐peptide hydrolyzing like activity: (a) positive cooperativity disappears as indicated by a lack of sigmoid initial parts of the kinetic curves, (b) substrate affinity is increased, whereby (c), the maximal activity remains virtually constant. As these kinetic features are independent of the peptide substrates, we conclude that PA28 exerts its activating influence on the proteasome by enhancing the uptake (and release) of shorter peptides.Keywords
This publication has 30 references indexed in Scilit:
- The Proteasome Activator 11 S Regulator or PA28Published by Elsevier BV ,1998
- Double‐cleavage production of the CTL epitope by proteasomes and PA28: role of the flanking regionGenes to Cells, 1997
- Structural and functional effects of PA700 and modulator protein on proteasomes 1 1Edited by W. BaumeisterJournal of Molecular Biology, 1997
- Relative Functions of the α and β Subunits of the Proteasome Activator, PA28Journal of Biological Chemistry, 1997
- Molecular cloning of the mouse proteasome subunits MC14 and MECL‐1: reciprocally regulated tissue expression of interferon‐γ‐modulated proteasome subunitsEuropean Journal of Immunology, 1997
- Structure of 20S proteasome from yeast at 2.4Å resolutionNature, 1997
- Molecular properties of the proteasome activator PA28 family proteins and γ‐interferon regulationGenes to Cells, 1997
- Reconstitution of proteasome activator PA28 from isolated subunits: optimal activity is associated with an α,β‐heteromultimerFEBS Letters, 1996
- A role for the proteasome regulator PA28α in antigen presentationNature, 1996
- Incorporation of major histocompatibility complex – encoded subunits LMP2 and LMP7 changes the quality of the 20S proteasome polypeptide processing products independent of interferon‐γEuropean Journal of Immunology, 1995