Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Dec 31;23(1):443.
doi: 10.3390/ijms23010443.

Oxidized Substrates of APEH as a Tool to Study the Endoprotease Activity of the Enzyme

Annamaria Sandomenico  1 , Marta Gogliettino  2 , Emanuela Iaccarino  1 , Carmela Fusco  2   3 , Andrea Caporale  1 , Menotti Ruvo  1 , Gianna Palmieri  2 , Ennio Cocca  2
Affiliations

Oxidized Substrates of APEH as a Tool to Study the Endoprotease Activity of the Enzyme

Annamaria Sandomenico et al. Int J Mol Sci. .

Abstract

APEH is a ubiquitous and cytosolic serine protease belonging to the prolyl oligopeptidase (POP) family, playing a critical role in the processes of degradation of proteins through both exo- and endopeptidase events. Endopeptidase activity has been associated with protein oxidation; however, the actual mechanisms have yet to be elucidated. We show that a synthetic fragment of GDF11 spanning the region 48-64 acquires sensitivity to the endopeptidase activity of APEH only when the methionines are transformed into the corresponding sulphoxide derivatives. The data suggest that the presence of sulphoxide-modified methionines is an important prerequisite for the substrates to be processed by APEH and that the residue is crucial for switching the enzyme activity from exo- to endoprotease. The cleavage occurs on residues placed on the C-terminal side of Met(O), with an efficiency depending on the methionine adjacent residues, which thereby may play a crucial role in driving and modulating APEH endoprotease activity.

Keywords: APEH; endoproteolytic activity; oxidative stress; oxidized methionine; oxidized substrates.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Scheme of the hydrolysis of GDF11[48-64]ox by trypsin and identification of the oxidation sites on methionines.
Figure 2
Figure 2
LC-MS analysis of GDF11[48-64]ox following tryptic digestion. Base peaks profiles of the peptide after oxidation and treatment with trypsin (A). EIC analysis of Fragment 1 (Ac-EYMFMQK), (B). EIC analysis of Fragment 2 (YPHTHLVQQA-NH2), (C). Mass spectrum of Fragment 1, (D). Mass spectrum of Fragment 2, (E).
Figure 3
Figure 3
LC-MS analysis of the GDF11[48-64]ox before and after treatment with APEH for 48 h. Base Peak profiles of GDF11[48-64]ox (A) before and (B) after treatment with APEH where the 3 peaks generated are indicated by arrows. (C–F) EIC profiles of Peaks 1–3. (F–I) Mass spectra of Peak 1, Peak 2 and Peak 3. Under Peak 3, two main species were identified as the fragments Ac-EYM(ox)FM(ox)QKYPHTHLVQ (species c) and Ac-EYM(ox)FM(ox)QKYPHTHL (species d). The species identified are reported on the mass spectra.
Figure 4
Figure 4
Scheme of main cleavage site of APEH on GDF11[48-64]ox.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Valko M., Leibfritz D., Moncol J., Cronin M.T., Mazur M., Telser J. Free radicals and antioxidants in normal physiological functions and human disease. Int. J. Biochem. Cell Biol. 2007;39:44–84. doi: 10.1016/j.biocel.2006.07.001. - DOI - PubMed
    1. Garcia-Rojo G., Gamiz F., Ampuero E., Rojas-Espina D., Sandoval R., Rozas C., Morales B., Wyneken U., Pancetti F. In Vivo Sub-chronic Treatment with Dichlorvos in Young Rats Promotes Synaptic Plasticity and Learning by a Mechanism that Involves Acylpeptide Hydrolase Instead of Acetylcholinesterase Inhibition. Correlation with Endogenous beta-Amyloid Levels. Front. Pharmacol. 2017;8:483. doi: 10.3389/fphar.2017.00483. - DOI - PMC - PubMed
    1. Scaloni A., Jones W.M., Barra D., Pospischil M., Sassa S., Popowicz A., Manning L.R., Schneewind O., Manning J.M. Acylpeptide hydrolase: Inhibitors and some active site residues of the human enzyme. J. Biol. Chem. 1992;267:3811–3818. doi: 10.1016/S0021-9258(19)50598-1. - DOI - PubMed
    1. Fujino T., Tada T., Beppu M., Kikugawa K. Purification and characterization of a serine protease in erythrocyte cytosol that is adherent to oxidized membranes and preferentially degrades proteins modified by oxidation and glycation. J. Biochem. 1998;124:1077–1085. doi: 10.1093/oxfordjournals.jbchem.a022224. - DOI - PubMed
    1. Shimizu K., Ikegami-Kawai M., Takahashi T. Increased oxidized protein hydrolase activity in serum and urine of diabetic rat models. Biol. Pharm. Bull. 2009;32:1632–1635. doi: 10.1248/bpb.32.1632. - DOI - PubMed

LinkOut - more resources