Extended cleavage specificity of a Chinese alligator granzyme B homologue, a strict Glu-ase in contrast to the mammalian Asp-ases

Ryu, Jinhye; Fu, Zhirong; Akula, Srinivas; Olsson, Anna-Karin; Hellman, Lars

doi:10.1016/j.dci.2021.104324

Cited by 7 publications

(5 citation statements)

References 27 publications

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“…This is based on its identification in a frog, with an active site that appears from in-silico analysis to have asp-ase or glu-ase activity [ 6 ]. A granzyme B homologue is also clearly present in reptiles, as seen in the Chinese alligator, as well as in monotremes, as exemplified by the platypus and also in a marsupial, the American opossum [ 14 , 15 , 20 ]. In these three species, there is also evidence for a classical mast cell chymase and/or a cathepsin G homologue [ 14 , 15 , 20 ].…”

Section: Discussionmentioning

confidence: 99%

“…A granzyme B homologue is also clearly present in reptiles, as seen in the Chinese alligator, as well as in monotremes, as exemplified by the platypus and also in a marsupial, the American opossum [ 14 , 15 , 20 ]. In these three species, there is also evidence for a classical mast cell chymase and/or a cathepsin G homologue [ 14 , 15 , 20 ]. An early ancestor of the neutrophil proteases, proteinase 3/N-elastase, also seems to appear in amphibians [ 4 ].…”

Section: Discussionmentioning

confidence: 99%

“…Human cytotoxic T cells and NK cells express and store several granzymes with asp-ase, tryptase, chymase, and meth-ase specificities, as exemplified by granzymes B, A, K, H, and M, respectively [ 2 ]. A relatively detailed picture has also been established of how these enzymes have appeared and diversified during mammalian evolution [ 3 , 4 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 ]; however, the situation in reptiles, amphibians, and fish is still only very fragmentary [ 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 ].…”

Section: Introductionmentioning

confidence: 99%

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The Extended Cleavage Specificity of Channel Catfish Granzyme-like II, A Highly Specific Elastase, Expressed by Natural Killer-like Cells

Thorpe,

Akula,

et al. 2023

IJMS

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The extended cleavage specificity of catfish granzyme-like II has been characterized using substrate phage display. The preference for particular amino acids at and surrounding the cleavage site was further validated by using a panel of recombinant substrates. This serine protease, which has previously been isolated as cDNA from a catfish natural killer-like cell line showed a preference for Ala in the P1 position of the substrate, and for multiple basic amino acids N-terminally of the cleavage site. A closely related zebrafish serine protease (zebrafish esterase-like) showed a very similar cleavage specificity, indicating an evolutionary conservation of this protease specificity among various fish species. Two catfish serine proteases, originating from NK-like cells, have now been isolated and characterized. One of them is highly specific met-ase with similar characteristics as the mammalian granzyme M. This enzyme may be involved in the induction of apoptosis in virus-infected cells, with a potential target in (catfish) caspase 6. In contrast to catfish granzyme-like I, the second enzyme analyzed here does not seem to have a direct counterpart in mammalian NK cells, and its role in the immune function of catfish NK cells is, therefore, still not known. However, this enzyme seems to be able to cleave a number of cytoskeletal proteins, indicating a separate strategy to induce apoptosis in target cells. Both of these enzymes are very interesting targets for further studies of their roles in catfish immunity, as enzymes with similar specificities have also been identified in zebrafish.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The Extended Cleavage Specificity of Channel Catfish Granzyme-like II, A Highly Specific Elastase, Expressed by Natural Killer-like Cells

Thorpe,

Akula,

et al. 2023

IJMS

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“…A relatively detailed picture has been established of how these enzymes have appeared and diversified during mammalian evolution [2,3,[5][6][7][8][9][10][11][12][13][14][15]. However, the information concerning the presence and specificity of such enzymes in reptiles, amphibians, and fishes is still only fragmentary [16][17][18][19][20][21][22][23]. There has been an increase in the use of non-rodent animal models with much focus on nonmammalian vertebrates including fish species, notably the zebrafish.…”

Section: Introductionmentioning

confidence: 99%

Extended Cleavage Specificity of two Hematopoietic Serine Proteases from a Ray-Finned Fish, the Spotted Gar (Lepisosteus oculatus)

Valentini,

Akula,

Alvarado-Vazquez

et al. 2024

IJMS

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The extended cleavage specificities of two hematopoietic serine proteases originating from the ray-finned fish, the spotted gar (Lepisosteus oculatus), have been characterized using substrate phage display. The preference for particular amino acids at and surrounding the cleavage site was further validated using a panel of recombinant substrates. For one of the enzymes, the gar granzyme G, a strict preference for the aromatic amino acid Tyr was observed at the cleavable P1 position. Using a set of recombinant substrates showed that the gar granzyme G had a high selectivity for Tyr but a lower activity for cleaving after Phe but not after Trp. Instead, the second enzyme, gar DDN1, showed a high preference for Leu in the P1 position of substrates. This latter enzyme also showed a high preference for Pro in the P2 position and Arg in both P4 and P5 positions. The selectivity for the two Arg residues in positions P4 and P5 suggests a highly specific substrate selectivity of this enzyme. The screening of the gar proteome with the consensus sequences obtained by substrate phage display for these two proteases resulted in a very diverse set of potential targets. Due to this diversity, a clear candidate for a specific immune function of these two enzymes cannot yet be identified. Antisera developed against the recombinant gar enzymes were used to study their tissue distribution. Tissue sections from juvenile fish showed the expression of both proteases in cells in Peyer’s patch-like structures in the intestinal region, indicating they may be expressed in T or NK cells. However, due to the lack of antibodies to specific surface markers in the gar, it has not been possible to specify the exact cellular origin. A marked difference in abundance was observed for the two proteases where gar DDN1 was expressed at higher levels than gar granzyme G. However, both appear to be expressed in the same or similar cells, having a lymphocyte-like appearance.

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“…The best characterized of them is gzm B, which is involved in both caspase-dependent and independent activation of apoptosis in virus-infected cells (5)(6)(7). The gene coding for gzm B (or a gzm B-like protease) is positioned in the chymase locus and is found in all tetrapods, from amphibians to humans (8)(9)(10)(11). Gzm B, has Asp-ase activity, and the human chymase locus also includes gzm H, which has chymotryptic cleavage specificity (8,12).…”

Section: Introductionmentioning

confidence: 99%

Extended cleavage specificities of human granzymes A and K, two closely related enzymes with conserved but still poorly defined functions in T and NK cell-mediated immunity

Aybay,

Ryu,

et al. 2023

Front. Immunol.

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Granzymes A and K are two highly homologous serine proteases expressed by mammalian cytotoxic T cells (CTL) and natural killer cells (NK). Granzyme A is the most abundant of the different granzymes (gzms) expressed by these two cell types. Gzms A and K are found in all jawed vertebrates and are the most well conserved of all hematopoietic serine proteases. Their potential functions have been studied extensively for many years, however, without clear conclusions. Gzm A was for many years thought to serve as a key component in the defense against viral infection by the induction of apoptosis in virus-infected cells, similar to gzm B. However, later studies have questioned this role and instead indicated that gzm A may act as a potent inducer of inflammatory cytokines and chemokines. Gzms A and K form clearly separate branches in a phylogenetic tree indicating separate functions. Transcriptional analyses presented here demonstrate the presence of gzm A and K transcripts in both CD4+ and CD8+ T cells. To enable screening for their primary biological targets we have made a detailed analysis of their extended cleavage specificities. Phage display analysis of the cleavage specificity of the recombinant enzymes showed that both gzms A and K are strict tryptases with high selectivity for Arg over Lys in the P1 position. The major differences in the specificities of these two enzymes are located N-terminally of the cleavage site, where gzm A prefers small amino acids such as Gly in the P3 position and shows a relatively relaxed selectivity in the P2 position. In contrast, gzm K prefers large amino acids such as Phe, Tyr, and Trp in both the P2 and P3 positions and does not tolerate negatively charged residues in the P2 position. This major distinction in extended specificities is likely reflected also in preferred in vivo targets of these two enzymes. This information can now be utilized for high-precision screening of primary targets for gzms A and K in search of their highly conserved but still poorly defined functions in vertebrate immunity.

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Extended cleavage specificity of a Chinese alligator granzyme B homologue, a strict Glu-ase in contrast to the mammalian Asp-ases

Cited by 7 publications

References 27 publications

The Extended Cleavage Specificity of Channel Catfish Granzyme-like II, A Highly Specific Elastase, Expressed by Natural Killer-like Cells

The Extended Cleavage Specificity of Channel Catfish Granzyme-like II, A Highly Specific Elastase, Expressed by Natural Killer-like Cells

Extended Cleavage Specificity of two Hematopoietic Serine Proteases from a Ray-Finned Fish, the Spotted Gar (Lepisosteus oculatus)

Extended cleavage specificities of human granzymes A and K, two closely related enzymes with conserved but still poorly defined functions in T and NK cell-mediated immunity

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