1.C.39 The Membrane Attack Complex/Perforin (MACPF) Family
Complement is part of the mammalian immune defense system against pathogenic microorganisms. More than 20 complement proteins control microbial invasion. One mechanism involves direct killing of the microbe, dependent on the C5b-8-9 or membrane attack complex (MAC) (Wang et al., 2000). This complex forms on the outer surface of a Gram-negative bacterial cell. The C5b-8 complex probably allows entry of C9 into the periplasm where the inactive protoxin is converted to the active toxin. This conversion process probably involves sulfhydryl oxidation to a disulfide linked protein. It has been shown that (C5b-8)1 (C9)1 which can lyse erythrocytes can not kill bacteria, but a (C5b-8)1 (C9)4 complex is bactericidal and also protozoan pathogens (Wang et al., 2000). Only the C-terminal portion of C9, residues 145-538 (the C9b fragment) is required to dissipate the membrane potential across respiring inner membrane vesicles, and only C9 is required if the protein is first targeted to the periplasm by any mechanism. While C9 therefore appears to form a pore in the inner membrane, C5b-8 appears to allow access of C9 to the periplasm. C6, C7, C8α, C8β and C9 all contain the pore forming MACPF domain (Rosado et al., 2008).
Native complement protein C9 is a soluble glycoprotein devoid of cellular toxicity, but its bactericidal activity is unmasked in the periplasm. This requires disulfide bond formation, a presumed conformational change, and insertion of a 'processed' toxin into the inner bacterial membrane, probably as an oligomer. Four disulfide bonds can potentially form in C9, three in the epidermal growth factor domain (residues 487-518) and one in the membrane attack/perforin region (Cys359 and Cys384). C9 has a 3-dimensional fold for its membrane interaction and pore forming domain as many other toxins of animal and bacterial origin (Anderluh and Lakey, 2008; Lukoyanova and Saibil, 2008). Kurschus et al., 2008 provided evidence that perforins and cholesterol-dependent cytolysins can deliver deadly proteins such as granzymes into the cytoplasm of target cells.
Proteins of the complement membrane attack complex (MAC) and the protein perforin (PF) share a common MACPF domain that is responsible for membrane insertion and pore formation (Cajnko et al. 2014). Hadders et al. 2007 determined the crystal structure of the MACPF domain of complement component C8α at 2.5 angstrom resolution and showed that it is structurally homologous to the bacterial, pore-forming, cholesterol-dependent cytolysins (TC #1.C.12). The structure displays two regions that (in the bacterial cytolysins) refold into transmembrane ß hairpins, forming the lining of a barrel pore. Local hydrophobicity explains why C8α is the first complement protein to insert into the membrane. The size of the MACPF domain is consistent with known C9 pore sizes. Thus, these mammalian and bacterial cytolytic proteins share a common mechanism of membrane insertion (Hadders et al., 2007). Insertion entails a dramatic refolding of the protein structure (Lukoyanova et al. 2016).
Proteins containing membrane attack complex/perforin (MACPF) domains play important roles in vertebrate immunity, embryonic development, and neural-cell migration among others (Anderluh and Lakey, 2008). In vertebrates, the ninth component of complement and perforin form oligomeric pores that lyse bacteria and kill virus-infected cells, respectively. Rosada et al. 2007 determined the crystal structure of a bacterial MACPF protein, Plu-MACPF from Photorhabdus luminescens, to 2.0 angstrom resolution. The MACPF domain revealed structural similarity with pore-forming cholesterol-dependent cytolysins (CDCs; 1.C.12) from Gram-positive bacteria. This suggests that lytic MACPF proteins may use a CDC-like mechanism to form pores and disrupt cell membranes. Sequence similarity between bacterial and vertebrate MACPF domains suggests that the fold of the CDCs, a family of proteins important for bacterial pathogenesis, is probably used by vertebrates for defense against infection.
The secretory granule-mediated cell death pathway is the key mechanism for elimination of virus-infected and transformed target cells by cytotoxic lymphocytes (Voskoboinik et al. 2010). The formation of the immunological synapse between an effector and a target cell leads to exocytic trafficking of the secretory granules and the release of their contents, which include pro-apoptotic serine proteases, granzymes, and pore-forming perforin into the synapse. There, perforin polymerizes and forms a transmembrane pore that allows the delivery of granzymes into the cytosol where they initiate various apoptotic death pathways. Unlike relatively redundant individual granzymes, functional perforin is absolutely essential for cytotoxic lymphocyte function and immune regulation in the host. Perforin's structure and function as well as its role in immune-mediated diseases have been reviewed (Voskoboinik et al. 2010). pH influsences perforin's permeabilizing activity but not its binding to membranes (Praper et al. 2010).
Law et al. (2010) elucidated the mechanism of perforin pore formation by determining the X-ray crystal structure of monomeric murine perforin, together with a cryo-electron microscopy reconstruction of the entire perforin pore. Perforin is a thin 'key-shaped' molecule, comprising an amino-terminal membrane attack complex perforin-like (MACPF)/cholesterol dependent cytolysin (CDC) domain followed by an epidermal growth factor (EGF) domain that, together with the extreme carboxy-terminal sequence, forms a central shelf-like structure. A C-terminal C2 domain mediates initial, Ca2+-dependent membrane binding. Electron microscopy revealed that the orientation of the perforin MACPF domain in the pore is inside-out relative to the subunit arrangement in CDCs. These data reveal remarkable flexibility in the mechanism of action of the conserved MACPF/CDC fold (Law et al., 2010). The 2.5Å structure of human C8 protein provided mechanistic insight into membrane pore formation by complement. C8-C9 and C9-C9 interactions facilitate refolding and insertion of putative MACPF transmembrane β-hairpins to form a circular pore (Lovelace et al., 2011). Perforin's helices function as TMSs after membrane insertion (Neely et al. 2016).
Perforin (PFN) is produced by cytotoxic lymphocytes and aids in the clearance of tumor or virus-infected cells by a pore forming mechanism. Praper et al. (2011) showed that perforin forms heterogeneous pores with a broad range of conductances, from 0.15 to 21 nanosiemens. In comparison with large pores that possessed low noise and remained stably open, small pores exhibited high noise and were unstable. Furthermore, the opening step and the pore size were dependent on the lipid composition of the membrane. The heterogeneity in pore sizes was confirmed with cryo-electron microscopy and showed a range of sizes matching that observed in the conductance measurements. Two different membrane-bound PFN conformations were interpreted as pre-pore and pore states of the protein. PFN probably forms heterogeneous pores through a multistep mechanism.
The complement Membrane Attack Complex (MAC) forms transmembrane pores in pathogen membranes. The first step in MAC assembly is cleavage of C5 to generate metastable C5b, which forms a stable complex with C6, termed C5b-6. C5b-6 initiates pore formation via the sequential recruitment of homologous proteins: C7, C8, and 12-18 copies of C9, each of which comprise a central MACPF domain flanked by auxiliary domains. Aleshin et al. (2012) proposed a model of pore assembly, in which the auxiliary domains play key roles, both in stabilizing the closed conformation of the protomers, and in driving the sequential opening of the MACPF β-sheet of each new recruit to the growing pore. They also described an atomic model of C5b-6 at 4.2 Å resolution and showed that C5b provides 4 interfaces for the auxiliary domains of C6. The largest interface is created by the insertion of an interdomain linker from C6 into a hydrophobic groove created by a major reorganization of the α-helical domain of C5b. In combination with a rigid-body docking of N-terminal elements of both proteins, C5b becomes locked into a stable conformation. Both C6 auxiliary domains flanking the linker pack tightly against C5b. The net effect is to induce a clockwise rigid-body rotation of 4 auxiliary domains as well as an opening/twisting of the central β-sheet of C6, in the directions predicted to activate or prime C6 for the subsequent steps in MAC assembly. The complex also suggests novel small-molecule strategies for modulating pathological MAC assembly (Aleshin et al., 2012).
Cytotoxic lymphocytes use perforin to eliminate dangerous cells, while remaining refractory to lysis. At least two mechanisms jointly preserve the killer cell: the C-terminal residues of perforin dictate its rapid export from the ER, whose milieu otherwise favours pore formation. Perforin is then stored in secretory granules whose acidity prevent its oligomerisation. Following exocytosis, perforin delivers the proapoptotic protease, granzyme B, into the target cell by disrupting its plasma membrane. The defined crystal structure of the perforin monomer and cryo-electron microscopy (EM) of the entire pore suggest that passive transmembrane granzyme diffusion is the dominant proapoptotic mechanism (Lopez et al., 2012).
The membrane attack complex of complement (MAC), apart from its classical role of lysing cells, can also trigger a range of non-lethal effects on cells, acting to promote inflammation. Triantafilou et al. 2013 investigated these non-lethal effects on inflammasome activation and found that, following sublytic MAC attack, there is an increased cytosolic Ca2+ concentration, at least partly through Ca2+ release from the endoplasmic reticulum lumen via the inositol 1,4,5-triphosphate receptor (IP3R) and ryanodine receptor (RyR) channels (TC# 1.A.3). This increase in intracellular Ca2+ concentration leads to Ca2+ accumulation in the mitochondrial matrix via the 'mitochondrial calcium uniporter' (MCU; TC#1.A.77), and loss of mitochondrial transmembrane potential, triggering NLRP3 inflammasome activation and IL-1beta release. NLRP3 co-localises with the mitochondria, probably sensing the increase in calcium and the resultant mitochondrial dysfunction, leading to caspase activation and apoptosis (Triantafilou et al. 2013).
Proteins with MACPF domains have a variety of biological roles, including defence and attack, organism development, and cell adhesion and signalling (Ota et al. 2014). The distribution of these proteins in fungi appears to be restricted to some Pezizomycotina and Basidiomycota species only, in correlation with another group of proteins with unknown biological function, known as aegerolysins. These two protein groups coincide in only a few species, and they might operate in concert as cytolytic bi-component pore- forming agents. Representative proteins here include pleurotolysin B, which has a MACPF domain, and the aegerolysin-like protein pleurotolysin A, and the very similar ostreolysin A, which have been purified from oyster mushroom (Pleurotus ostreatus). These act in concert to perforate natural and artificial lipid membranes with high cholesterol and sphingomyelin content. The aegerolysin-like proteins provide the membrane cholesterol/sphingomyelin selectivity and recruit oligomerized pleurotolysin B molecules, to create a membrane-inserted pore complex. The resulting protein structure has been imaged with electron microscopy, and it has a 13-meric rosette-like structure, with a central lumen that is ~4-5 nm in diameter. The opened transmembrane pore is non-selectively permeable for ions and smaller neutral solutes and is a cause of cytolysis of a colloid-osmotic type.
D'Angelo et al. (2012) (PMID 22551122) identified orthologs and homologs of human perforin in all but one species analysed from Euteleostomi, and presented evidence for an earlier ortholog in Gnathostomata but not in more primitive chordates. In placental mammals, perforin is a single copy gene, but there are multiple perforin genes in all lineages predating marsupials, except birds. Comparisons of these many-to-one homologs of human perforin showed that they mainly arose from lineage-specific gene duplications in multiple taxa, suggesting acquisition of new roles or different modes of regulation. Perforins probably arose by duplication of the ancient MPEG1 gene (TC# 3.C.39.14), and that they share a common ancestor with the functionally related complement proteins. The evolution of perforin in vertebrates involved a complex pattern of gene/intron, gain and loss. The primordial perforin gene arose at least 500 million years ago, at around the time that the major histocompatibility complex-T cell receptor antigen recognition system was established. As it is absent from primitive chordates and invertebrates, cytotoxic cells from these lineages must possess a different effector molecule or cytotoxic mechanism.
CDC/MACPF proteins contain a characteristic four-stranded beta-sheet that is flanked by two alpha-helical bundles, which unfold to form two transmembrane beta-hairpins. Apicomplexan parasites express MACPFs termed perforin-like proteins (PLPs). Wade and Tweten 2015 present insights into the assembly and regulation of the Apicomplexan PLP molecular pore-forming mechanisms, necessary for osmotically driven rupture of the parasitophorous vacuole and host cell membrane, and cell traversal by these parasites.
Membrane Attack Complex (MAC) pores are assembled when surface-bound convertase enzymes convert C5 into C5b, which together with C6, C7, C8 and multiple copies of C9 forms transmembrane pores that damage the bacterial cell envelope. Bacterial killing requires local conversion of C5 by surface-bound convertases, and Doorduijn et al. 2020 showed that rapid interaction of C7 with C5b6 is required to form bactericidal MAC pores. This rapid interaction is crucial to efficiently anchor C5b-7 to the bacterial cell envelope. Bacterial pathogens can prevent complement-dependent killing by interfering with the anchoring of C5b-7 and thus prevented stable insertion of MAC pores into the bacterial cell envelope.
Unregulated complement activation causes inflammatory and immunological pathologies. To prevent damage during an immune response, extracellular chaperones (clusterin and vitronectin) capture and clear soluble precursors to the membrane attack complex (sMAC). Menny et al. 2021 combined cryoEM and cross-linking mass spectrometry (XL-MS) to solve the structure of sMAC. Clusterin recognizes and inhibits polymerizing complement proteins by binding a negatively charged surface of sMAC. The pore-forming C9 protein is trapped in an intermediate conformation whereby only one of its two transmembrane beta-hairpins has unfurled. This structure provides molecular details for immune pore formation and helps explain a complement control mechanism that has potential implications for how cell clearance pathways mediate immune homeostasis (Menny et al. 2021).
The generalized transport reaction catalyzed by MACPF family members is:
Small and large molecules (in) ⇌ small and large molecules (out)
Uncharacterized MACPF homologue of 1153 aas
UP of Dictyostelium fasciculatum (Slime mold)
Uncharacterized protein of 1277 aas
UP of Polysphondylium pallidum (Cellular slime mold)
Uncharacterized protein of 1216 aas
UP of Acytostelium subglobosum
Uncharacterized protein of 1151 aas
UP of Polysphondylium pallidum
Sea anemone toxin, AvTX-60A, of 498aas (Oshiro et al., 2004).
AvTX-60A of Actineria villosa (Q76DT2)
MACPF-containing actinoporin of 488 aas, PsTX60B (Frazão et al. 2012).
PsTX60B of Phyllodiscus semoni (Night anemone)
Uncharacterized protein of 449 aas
UP of Selaginella moellendorffii (Spikemoss)
Uncharacterized protein of 474 aas
UP of Nematostella vectensis (Starlet sea anemone)
MACPF protein (610aas)
MACPF protein of Medicago truncatula (Q1SKW8)
MACPF protein (615aas)
MACPF protein of Populus trichocarpa (B9GNC9)
The constitutively activated cell death 1 protein (CAD1) of 561 aas (Morita-Yamamuro et al. 2005; Tsutsui et al. 2006).
CAD1 of Arabidopsis thaliana
The Necrotic Spotted Lesions 1 (NSL1) protein of 612 aas (Noutoshi et al. 2006).
NSL1 of Arabidopsis thaliana
MACPF protein (809aas)
MACPF protein of Chlamydia muridarum (Q9PKN4)
MACPF homologue (411aas)
MACPF homologue of Chlamydophila pneumoniae (Q9Z908)
MACPF protein, CT153 of 810 aas. Mediates interactions with host cell membranes and organelles, and plays a role in intracellular development (Taylor and Nelson 2014).
CT153 protein of Chlamydia trachomatis
MACPF domain protein of 834 aas
MACPF protein of Chlamydia psittaci
Hypothetical protein (470aas)
HP of Bacteroides thetaiotaomicron (Q8A335)
MACPF-domain containing protein, BSAP-1, of 372 aas and 1 N-terminal TMS, secreted in extracellular vesicles. It contains a membrane attack complex/perforin (MACPF) domain that kills bacteria by pore formation, and mutations affecting key residues of this domain abrogated its activity (Chatzidaki-Livanis et al. 2014). Extracellular Vesicles can be relevant to Endocrinology in mammals (Das Gupta et al. 2021).
BSAP1 of Bacteroides fragilis (Q64VU4)
Hypothetical Protein (486 aas)
HP of Bacteroides fragilis (Q64W10)
MACPF protein. The structure is known (Xu et al., 2010).
MACPF protein of Bacteroides thetaiotaomicron (Q8A267)
UP of Paraprevotella xylaniphila
MAC/Perforin domain protein BSAP-4 of 506 aas and 1 N-terminal TMS.
BSAP-4 of Bacteroides fragilis
MACPF domain-containing protein, BASP2, of 508 aas (Roelofs et al. 2016).
BASP2 of Bacteroides uniformis
MACPF domain-containing protein, BSAP3, of 485 aas and one N-terminal TMS (McEneany et al. 2018).
BSAP3 of Bacteroides dorei
Macrophage-expressed gene 1 protein, Mpeg1, or perforin-2, PFN2, of 716 aas and one C-terminal TMS. Pore formation has been demonstrated in target bacteria (McCormack et al. 2013). PFN2 undergoes a pre-pore to pore transition upon acidification (Jiao et al. 2021).
Mpeg1 of Homo sapiens
MACPF domain protein, Mpeg of 742 aas. This protein is found in late endosomes. Its MACPF domain exhibits anti-bacterial activity against Gram - and Gram + bacteria. It's synthesis is stimulated following infection with Vibrio alginolyticus (He et al. 2011).
Mpeg of Crassostrea gigas (Pacific oyster) (Crassostrea angulata)
Mpeg1 of 728 aas. Contains a cytolytic MACPF domain. Expressed in up to 8x increase in hematocytes and epipodia samples after exposure to heat killed Vibrio anguilarum (Kemp and Coyne 2011).
Mpeg1 of Haliotis midae (perlemoen abalone)
Mpeg1 of 718 aas (Benard et al. 2014).
Mpeg1 of Danio rerio (Zebrafish) (Brachydanio rerio)
Torso-like protein, Tsl of 353 aas and containing a MACPF domain. Possible ligand that binds to the torso receptor. Implicated in a receptor tyrosine kinase signaling pathway that specifies differentialtion and terminal cell fate (Martin et al. 1994; Savant-Bhonsale and Montell 1993; Johnson et al. 2013; Mineo et al. 2015).
Tsl of Drosophila melanogaster (Fruit fly)
Uncharacterized Torso-like protein of 271 aas
Torso-like protein of Daphnia pulex (Water flea)
Uncharacterized protein of 784 aas
UP of Penicillium marneffei
Uncharacterized protein of 795 aas
UP of Cladophialophora psammophila
The BMP/retinoic acid-inducible neural-specific protein 1, BRINP1 (DBC1, DBCCR1, FAM5A), protein of 761 aas. Inhibits cell proliferation by negative regulation of the G1/S transition and mediates cell death which is not of the classical apoptotic type while regulating expression of components of the plasminogen pathway (Wright et al. 2004; Nishiyama et al. 2001; Louhelainen et al. 2006).
BRINP-1 of Homo sapiens
BRINP-2 (BRINP2, FAM5B) of 783 aas. Inhibits neuronal cell proliferation by negative regulation of the cell cycle transition.
BRINP-2 of Homo sapiens
Perforin 1 precursor; targets viruses, bacteria and cancer cells (McCormack et al. 2013). It is produced by cytotoxic T lymphocytes and natural killer cells and has been expressed, purified and studied in insect cells (Naneh et al. 2015).
Perforin of Rattus norvegicus
Uncharacterized protein of 429 aas
UP of Astyanax mexicanus (Blind cave fish) (Astyanax fasciatus mexicanus)
Performin 1-like protein of 545 aas, Prf1
Prf1 of Xenopus tropicalis (Western clawed frog) (Silurana tropicalis)
Uncharacterized protein of 481 aas
UP of Latimeria chalumnae (West Indian ocean coelacanth)
Performin 1, Pfn1, of 587 aa
Pfn1 of Carassius auratus langsdorfii (Japanese silver crucian carp)
MACPF domain-containing protein (342aas)
MACPF proteins of Tetrahymena thermophila (Q23MJ4)
Duplicated MACPF protein (681aas) The first half resembles 1.C.39.6.2 more than the second half.
MACPF protein of Tetrahymena thermophila (Q23I78)
Perforin 1, PRF1 of 555 aas. Plays a key role in secretory granule-dependent cell death and in defense against virus-infected and neoplastic cells. Plays an important role in killing other cells that are recognized as non-self by the immune system, e.g. in transplant rejection or some forms of autoimmune disease. Can insert into the membrane of target cells in its calcium-bound form, oligomerize and form large pores (Law et al. 2010). Promotes cytolysis and apoptosis of target cells by facilitating the uptake of cytotoxic granzymes. Perforin gene mutations contribute to hereditary cancer predisposition (Chaudhry et al. 2016). After perforin is secreted by CD8+ cytotoxic T-lymphocytes (CTLs) and disrupts the membranes of extracellular vesicles (EVs), adenosine is released from the EVs and acts as an immunosuppressive metabolite by binding to the adenosine receptor on the CTL membrane. This mechanism provides a novel survival strategy using cancer cell-derived EVs (Tadokoro et al. 2020).
Perforin of Homo sapiens
Pore-forming, membrane attack, complement component 8, α-polypeptide precursor; C8α-MACPF (structure solved to 2.5 Å resolution; Hadders et al., 2007; Rosado et al., 2007). β-Hairpins in C8α and C9 play a direct role in MAC membrane penetration and pore formation (Weiland et al. 2014). The first TMS of complement component-9 inhibits its own self assembly (Spicer et al. 2018).
C8α-MACPF of Homo sapiens (2QQH_A) (P07357)
Complement component 7
Complement component 7 of Xenopus laevis (Q6INM0)
Complement component C6 of 934 aas and 1 TMS; targets phagocytic and some non phagocytic cells (McCormack et al. 2013). Expressed constitutively in phagocytes and inducibly in parenchymal tissue-forming cells. It is a transmembrane protein of cytosolic vesicles, derived from multiple organelles that translocate to and fuse with bacterium-containing vesicles. Subsequently, perforin-2 polymerizes and forms large clusters of 100 Å pores in the bacterial surface with perforin-2 cleavage products present in the bacteria. Perforin-2 is also required for the bactericidal activity of reactive oxygen and nitrogen species as well as hydrolytic enzymes (McCormack et al. 2015). Perforin-2 exists in membrane-bound (P2a) and secretory (P2b) isoforms, both present in human macrophages. P2a promotes fusion of vesicles with lysosomes, and may therefore play important roles in immune defense (Xiong et al. 2017). Loss of MPEG1 causes increased susceptibility to microbial infection. MPEG1 expression is upregulated in response to proinflammatory signals such as tumor necrosis factor alpha (TNFα) and lipopolysaccharides (LPS). Furthermore, germline mutations in MPEG1 have been identified in connection with recurrent pulmonary mycobacterial infections. Structural studies on MPEG1 revealed that it can form oligomeric pre-pores and pores. The unusual domain arrangement within the MPEG1 architecture suggests a novel mechanism of pore formation that may have evolved to guard against unwanted lysis of host cells (Bayly-Jones et al. 2020).
Complement component C of Homo sapiens
MACPF protein, terminal complement component, TCC-like of 585 aas.
TCC-like protein of Halocynthia roretzi (Sea squirt) (Cynthia roretzi)
Complement protein C9; targets bacteria (McCormack et al. 2013). β-Hairpins in C8α and C9 play a direct role in MAC membrane penetration and pore formation (Weiland et al. 2014).
C9 of Equus caballus
Complement protein C9. β-Hairpins in C8α and C9 play a direct role in MAC membrane penetration and pore formation (Weiland et al. 2014).
C9 of Fugu rubripes
Chain A, MACPF perforin-like protein, Plu-MACPF (structure solved to 2.0 Å resolution; Rosado et al., 2007).
Plu-MACPF of Photorhabdus luminescens (2QP2_A) (Q7N6X0)
MACPF protein (453 aas)
MACPF protein of Trichodesmium erythraeum (Q117U3)
Hypothetical Protein (588 aas)
HP of Marinomonas sp. MED121 (A3YG19)
Putative perforin of 409 aas
Putative perforin of Halorubrum kocurii
Uncharacterized protein of 684 aas
UP of Selaginella moellendorffii
Uncharacterized protein of 1085 aas. This protein is a fusion protein with an N-terminal MACPF domain (see TC subfamily # 1.C.39..4) and a C-terminal internalin-A domain (see TC# 8.A.43.1.12).
UP of Acholeplasma palmae
Gram-negative insecticidal protein, GNIP1Aa of 536 aas and 0 TMSs/ Its structure has been determined to 2.5 Å resolution (PDB# 6FBM) (Zaitseva et al. 2019). It consists of two structurally distinct domains, a MACPF domain and a previously uncharacterized type of domain. GNIP1Aa is unique in being a prokaryotic MACPF member to have both its structure and function identified. It is specifically toxic to Diabrotica virgifera virgifera larvae upon feeding. The MACPF domain is probably important for oligomerization and transmembrane pore formation, while the accompanying domain may define the specificity of the target of toxicity. In GNIP1Aa the accompanying C-terminal domain has a unique fold composed of three pseudosymmetric subdomains with shared sequence similarity, a feature not obvious from the initial sequence examination. This domain is in a family named beta-tripod. Important regions in the beta-tripod domain, which may be involved in target recognition, have been identified (Zaitseva et al. 2019).
GNIP1 of Chromobacterium piscinae
Uncharacterized phosphodiesterase of 771 aas and 0 TMSs. Only a segment of this protein is homologous to other members of family 1.C.39, and this segment is also distantly related to members of family 1.C.43.
UP of Scytonema sp. NIES-4073
Phosphodiesterase/alkaline phosphatase D, PhoD, of 837 aas and 0 TMSs. Except for the protein with TC# 1.C.39.4.8, only a segment of this protein is homologous to other members of the MACPF family, Iit is also related to members of family 1.C.43.
PhoD of Calothrix brevissima
Fish (Lancelets; Branchiostomidae)
MACPF homologue of Branchiostoma floridae (C3YI39)
Starlet Sea Anemone
MACPF homologue of Nematostella vectensis (A7RF41)
Fish (Lancelets; Branchiostomidae)
MACPF homologue of Branchiostoma floridae (C3Z435)
Protein of 1305 aas with an N-terminal MACPF domain and C-terminal extracellular cystine-rich furin-like (Fu-sup), fucolectin (tocylectin; discoidin; FTP; a fucose-binding lectin) and cystine-rich scavenger receptor (SRCR; extracellular protein-protein interaction) domains (in this order, N- to C-terminus).
MACPF protein of Branchiostoma floridae (Florida lancelet) (Amphioxus)
Cholesterol-dependent cytolysin of 632 aas
UP of Pseudomonas thivervalensis
Sporozoite protein with MAC/Perforin domain (Homologous to Erylysin B) of 810 aas. Interacts and breaches host cell membranes (Tavares et al. 2014). CDC/MACPF proteins contain a characteristic four-stranded beta-sheet that is flanked by two alpha-helical bundles, which unfold to form two transmembrane beta-hairpins. Apicomplexan parasites express CDC/MACPFs termed perforin-like proteins (PLPs). Wade and Tweten 2015 present insights into the assembly and regulation of the Apicomplexan CDC (ApiMACPF) molecular pore-forming mechanisms, necessary for osmotically driven rupture of the parasitophorous vacuole and host cell membrane, and cell traversal by these parasites.
MACPF protein of Plasmodium knowlesi (B3L016)
Perforin-like protein, PLP1, of 1150 aas (Tavares et al. 2014).
PLP1 of Toxoplasma gondii
MACPF protein of Theileria parva (Q4MYP3)
MACPF domain-containing protein (420aas)
MACPF protein of Babesia bovis (A7AT97)
MAC/Perforin domain protein
MACPF domain protein of Tetrahymena thermophila (Q23QV5)
Uncharacterized protein of 1040 aas
UP of Capsaspora owczarzaki
The MAC/Perforin domain containing protein of 861 aas
MACPF protein of Oxytricha trifallax
Apextrin of 853 aas
Apextrin of Acropora millepora (Staghorn coral)
Putative uncharacterized phospholipase D endonuclease of 487 aas
UP of Myxococcus fulvus
MACPF-Hemopexin protein. The MACPF domain forms pores in the membrane while the hemopexin domain fuctions as a heme scavenging domain, protecting the cell against heme toxicity (Mehta and Reddy 2015).
Hemopexin of Plesiocystis pacifica (A6G7F3)
The MACPF protein homologue with hemopexin-like C-terminal repeats
MACPF protein of Beggiotoa sp. PS (A7BVI9)
Photopexin a/b-like protein of 347 aas.
Photopexin of Photorhabdus temperat
MACPF homologue of Postia placenta (B8PKX3)
Uncharacterized MACPF protein of 446 aas. The MACPF domain includes residues 120 - 320.
UP of Emericella nidulans (Aspergillus nidulans)
Uncharacterized MACPF protein of 483 aas
UP of Fusarium oxysporum f. sp. vasinfectum
Uncharacterized MACPF protein of 420 aas
UP of Trichophyton verrucosum
Uncharacterized protein of 461 aas
UP of Ceriporiopsis subvermispora (White-rot fungus)