Supramolecular structure of tripeptidyl peptidase II from human erythrocytes as studied by electron microscopy, and its correlation to enzyme activity

Macpherson, E; Tomkinson, Birgitta; Bålöw, R M; Höglund, Stefan; Zetterqvist, Örjan

doi:10.1042/bj2480259

Cited by 47 publications

(35 citation statements)

References 11 publications

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“…7B). TPP II is a multimeric protease complex that may require proper assembly for normal function (26,48). Superose 6 gel filtration chromatography revealed that the increase in AAF-AMC hydrolyzing activity detected under these conditions of TPP II vac infection corresponded to a single peak eluting just before the 26S proteasome (Figs.…”

Section: Overexpression Of Tpp IImentioning

confidence: 97%

Re-evaluating the Generation of a “Proteasome-Independent” MHC Class I-Restricted CD8 T Cell Epitope

Wherry

Golovina

Morrison

et al. 2006

The Journal of Immunology

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The proteasome is primarily responsible for the generation of MHC class I-restricted CTL epitopes. However, some epitopes, such as NP147–155 of the influenza nucleoprotein (NP), are presented efficiently in the presence of proteasome inhibitors. The pathways used to generate such apparently “proteasome-independent” epitopes remain poorly defined. We have examined the generation of NP147–155 and a second proteasome-dependent NP epitope, NP50–57, using cells adapted to growth in the presence of proteasome inhibitors and also through protease overexpression. We observed that: 1) Ag processing and presentation proceeds in proteasome-inhibitor adapted cells but may become more dependent, at least in part, on nonproteasomal protease(s), 2) tripeptidyl peptidase II does not substitute for the proteasome in the generation of NP147–155, 3) overexpression of leucine aminopeptidase, thymet oligopeptidase, puromycin-sensitive aminopeptidase, and bleomycin hydrolase, has little impact on the processing and presentation of NP50–57 or NP147–155, and 4) proteasome-inhibitor treatment altered the specificity of substrate cleavage by the proteasome using cell-free digests favoring NP147–155 epitope preservation. Based on these results, we propose a central role for the proteasome in epitope generation even in the presence of proteasome inhibitors, although such inhibitors will likely alter cleavage patterns and may increase the dependence of the processing pathway on postproteasomal enzymes.

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Section: Overexpression Of Tpp IImentioning

confidence: 97%

Re-evaluating the Generation of a “Proteasome-Independent” MHC Class I-Restricted CD8 T Cell Epitope

Wherry

Golovina

Morrison

et al. 2006

The Journal of Immunology

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“…thimet oligopeptidase, prolyl oligopeptidase, aminopeptidases, or insulin-degrading enzyme) as determined with specific fluorogenic substrates or by immunoblotting. However, this step did remove the very high molecular weight enzyme, TPP-II (34,35). This enzyme has been proposed to play a role in final steps of intracellular protein degradation by degrading peptides generated by proteasomes (36).…”

Section: Antigenic Peptides Are Rapidly Digested By Cytosolicmentioning

confidence: 99%

Major Histocompatibility Complex Class I-presented Antigenic Peptides Are Degraded in Cytosolic Extracts Primarily by Thimet Oligopeptidase

Šarić¹,

Beninga²,

Graef³

et al. 2001

Journal of Biological Chemistry

136

106

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Nearly all peptides generated by proteasomes during protein degradation are digested rapidly to amino acids, but a few proteasomal products escape this fate and are presented to the immune system on cell surface major histocompatibility complex class I molecules. To test whether these antigenic peptides may be inherently resistant to cytosolic peptidases, six different antigenic peptides were incubated with HeLa cell extracts. All six were degraded rapidly by a process involving o-phenanthroline-sensitive metallopeptidases. One antigenic peptide, FAPGNYPAL, was rapidly destroyed in the extracts by a bestatin-sensitive exopeptidase, apparently by the puromycin-sensitive aminopeptidase. The disappearance of the other five was reduced 30 -90% by a specific inhibitor of the cytosolic endopeptidase, thimet oligopeptidase (TOP) (EC 3.4.24.15), whose physiological function(s) have been unclear and controversial. All these peptides were sensitive to pure recombinant TOP. Furthermore, upon fractionation of the extracts, the major peptidase peak that degraded the ovalbumin-derived epitope, SIINFEKL, co-purified with TOP. In the extracts, TOP also catalyzed rapid degradation of Nextended variants of SIINFEKL and of other antigenic peptides, which in vivo can serve as precursors of these major histocompatibility complex-presented epitopes. This enzyme (unlike cell proteins that promote production of antigenic peptides) is not regulated by interferon-␥. TOP seems to be primarily responsible for the rapid breakdown of antigenic peptides in cytosolic extracts, and our related studies

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“…In the latter case, the activity dropped to about one-tenth and recovered to some extent upon partial reassembly. This hinted at a correlation between oligomeric state and specific activity and elicited speculations on a regulation of the enzyme activity via association/dissociation (2,29,30).…”

mentioning

confidence: 98%

“…The TPP II holo-complex has been reported to decay under conditions such as dialysis (29) or freezing/thawing (30), with a concomitant decrease of activity. In the latter case, the activity dropped to about one-tenth and recovered to some extent upon partial reassembly.…”

mentioning

confidence: 99%

Size Matters for the Tripeptidylpeptidase II Complex from Drosophila

Seyit

Rockel

Baumeister

et al. 2006

Journal of Biological Chemistry

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Tripeptidylpeptidase II (TPP II) is an exopeptidase of the subtilisin type of serine proteases, a key component of the protein degradation cascade in many eukaryotes, which cleaves tripeptides from the N terminus of proteasome-released products. The Drosophila TPP II is a large homooligomeric complex (ϳ6 MDa) that is organized in a unique repetitive structure with two strands each composed of ten stacked homodimers; two strands intertwine to form a spindleshaped structure. We report a novel procedure of preparing an active, structurally homogeneous TPP II holo-complex overexpressed in Escherichia coli. Assembly studies revealed that the specific activity of TPP II increases with oligomer size, which in turn is strongly concentration-dependent. At a TPP II concentration such as prevailing in Drosophila, equilibration of size and activity proceeds on a time scale of hours and leads to spindle formation at a TPP II concentration of >0.03 mg/ml. Before equilibrium is reached, activation lags behind assembly, suggesting that activation occurs in a twostep process consisting of (i) assembly and (ii) a subsequent conformational change leading to a switch from basal to full activity. We propose a model consistent with the hyperbolic increase of activity with oligomer size. Spindle formation by strand pairing causes both significant thermodynamic and kinetic stabilization. The strands inherently heterogeneous in length are thus locked into a discrete oligomeric state. Our data indicate that the unique spindle form of the holo-complex represents an assembly motif stabilizing a highly active state.

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Supramolecular structure of tripeptidyl peptidase II from human erythrocytes as studied by electron microscopy, and its correlation to enzyme activity

Cited by 47 publications

References 11 publications

Re-evaluating the Generation of a “Proteasome-Independent” MHC Class I-Restricted CD8 T Cell Epitope

Re-evaluating the Generation of a “Proteasome-Independent” MHC Class I-Restricted CD8 T Cell Epitope

Major Histocompatibility Complex Class I-presented Antigenic Peptides Are Degraded in Cytosolic Extracts Primarily by Thimet Oligopeptidase

Size Matters for the Tripeptidylpeptidase II Complex from Drosophila

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