A novel class of bifunctional acylpeptide hydrolases – potential role in the antioxidant defense systems of the Antarctic fish Trematomus bernacchii

Abstract

Oxidative challenge is an important factor affecting the adaptive strategies of Antarctic fish, but data on antioxidant defenses in these organisms remain scarce. In this context, a key role could be played by acylpeptide hydrolase (APEH), which was recently hypothesized to participate in the degradation of oxidized and cytotoxic proteins, although its physiological function is still not fully clarified. This study represents the first report on piscine members of this enzyme family, specifically from the Antarctic teleost Trematomus bernacchii. The cDNAs corresponding to two apeh genes were isolated, and the respective proteins were functionally and structurally characterized with the aim of understanding the biological significance of these proteases in Antarctic fish. Both APEH isoforms (APEH-1 Tb and APEH-2 Tb ) showed distinct temperature-kinetic behavior, with significant differences in the K m values. Moreover, beside the typical acylpeptide hydrolase activity, APEH-2 Tb showed remarkable oxidized protein endohydrolase activity towards oxidized BSA, suggesting that this isoform could play a homeostatic role in removing oxidatively damaged proteins, sustaining the antioxidant defense systems. The 3D structures of both APEHs were predicted, and a possible relationship was found between the substrate specificity/affinity and the marked changes in the number of charged residues and hydrophobicity properties surrounding their catalytic sites. Our results demonstrated the occurrence of two APEH isoforms in T. bernacchii, belonging to different phylogenetic clusters, identified for the first time, and showing distinct molecular and temperature-kinetic behaviors. In addition, we suggest that the members of the new cluster 'APEH-2' could participate in reactive oxygen species detoxification as phase 3 antioxidant enzymes, enhancing the protein degradation machinery.

A New APEH Cluster with Antioxidant Functions in the Antarctic Hemoglobinless Icefish Chionodraco hamatus

“…Recently, acylpeptide hydrolase (APEH), which was first identified as oxidised protein hydrolase (OPH), has been hypothesised to participate in ROS detoxification as a member of the phase 3 antioxidant enzymes [ 11 ], which are involved in the elimination of irreversibly denatured proteins. Confirming this hypothesis, we showed in a previous study [ 12 ] that APEH plays a key role in the degradation of oxidised and cytotoxic proteins specifically in Antarctic fish.…”

“…We identified the first APEHs from the Antarctic fish Trematomus bernacchii [ 12 ]. Specifically, we demonstrated the presence of two active APEH isoforms (APEH-1 Tb and APEH-2 Tb ) belonging to different phylogenetic clusters and exhibiting distinct molecular and temperature–kinetic behaviours.…”

“…Specifically, we demonstrated the presence of two active APEH isoforms (APEH-1 Tb and APEH-2 Tb ) belonging to different phylogenetic clusters and exhibiting distinct molecular and temperature–kinetic behaviours. Interestingly, APEH-1 Tb showed the typical enzymatic properties of APEHs studied so far, whereas APEH-2 Tb appeared to be a bifunctional enzyme that exhibits exopeptidase activity towards N-acyl peptides and efficient endoprotease activity towards oxidised proteins, indicating that this isoform plays a key homeostatic role in sustaining the protective systems required to counteract cold-induced oxidative stress [ 12 ].…”

The loss of hemoglobin and myoglobin does not minimize oxidative stress in Antarctic icefishes

“…Studies of levels of chaperones in icefishes would address this question. More recently, a novel isoform of acylpeptide hydrolase (APEH-2) has been shown to effectively hydrolyze oxidized proteins (Gogliettino et al, 2014). The relative expression and activity of APEH-2 is higher in erythrocytes of the red-blooded notothenioid T. bernacchii than in the erythrocyte-like cells of the icefish Chionodraco hamatus, suggesting that this pathway may be important for eliminating oxidized proteins in red-blooded notothenioids (Riccio et al, 2015).…”

APEH Inhibition Affects Osteosarcoma Cell Viability via Downregulation of the Proteasome

“…APEH is one of the four members of the Prolyl OligoPeptidase class (POP, clan SC, family S9) [17]; it catalyzes the removal of N -acylated amino acids from acetylated peptides and it has been studied in a number of eukaryal [18], bacterial [19], and archaeal [20] organisms, although its physiological role has not been completely clarified. However, as previously suggested, APEH can be involved in the protein-degradation machinery, possibly acting as an antioxidant defense system [21,22,23,24,25], and could represent a promising therapeutic target for a wide array of diseases caused by misfolded protein accumulation [26,27]. …”