Kunitz domain of epididymal protease inhibitor eppin and similar proteins; This subfamily includes the Kunitz inhibitor domain protein eppin (also called Cancer/testis antigen 71 or CT71, epididymal protease inhibitor, protease inhibitor WAP7, serine protease inhibitor-like with Kunitz and WAP domains 1, or WAP four-disulfide core domain protein 7) as well as WAP four-disulfide core domain proteins 6A and 6B in mice, and similar proteins. Eppin is a serine protease inhibitor that plays an essential role in male reproduction and fertility. It modulates the hydrolysis of seminal fluid protein semenogelin 1 (SEMG1) by the serine protease kallikrein-related peptidase 3 (KLK3, PSA), provides antimicrobial protection for spermatozoa in the ejaculate coagulum, and binds SEMG1, thereby inhibiting sperm motility. Thus, eppin could potentially be used as a target for male contraception. These domains are similar to Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor) that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|....*..
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCKKKQ 188
Cdd:cd22611    1 DVCSLPKESGPCMAYFPRWWYDKETNTCSKFIYGGCQGNNNNFQSEAICQNICKKKR 57

Kunitz domain of epididymal protease inhibitor eppin and similar proteins; This subfamily includes the Kunitz inhibitor domain protein eppin (also called Cancer/testis antigen 71 or CT71, epididymal protease inhibitor, protease inhibitor WAP7, serine protease inhibitor-like with Kunitz and WAP domains 1, or WAP four-disulfide core domain protein 7) as well as WAP four-disulfide core domain proteins 6A and 6B in mice, and similar proteins. Eppin is a serine protease inhibitor that plays an essential role in male reproduction and fertility. It modulates the hydrolysis of seminal fluid protein semenogelin 1 (SEMG1) by the serine protease kallikrein-related peptidase 3 (KLK3, PSA), provides antimicrobial protection for spermatozoa in the ejaculate coagulum, and binds SEMG1, thereby inhibiting sperm motility. Thus, eppin could potentially be used as a target for male contraception. These domains are similar to Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor) that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|....*..
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCKKKQ 188
Cdd:cd22611    1 DVCSLPKESGPCMAYFPRWWYDKETNTCSKFIYGGCQGNNNNFQSEAICQNICKKKR 57

BPTI/Kunitz family of serine protease inhibitors; Serine protease inhibitors. One member of the family is encoded by an alternatively-spliced form of Alzheimer's amyloid beta-protein.

                           10        20        30        40        50
                   ....*....|....*....|....*....|....*....|....*....|...
gi 755499091   132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:smart00131   1 DVCLLPPDTGPCGGSIPRYYYDPETGTCEPFTYGGCGGNANNFESLEECERTC 53

whey acidic protein-type four-disulfide core domains. Members of the family include whey acidic protein, elafin (elastase-specific inhibitor), caltrin-like protein (a calcium transport inhibitor) and other extracellular proteinase inhibitors. A group of proteins containing 8 characteristically-spaced cysteine residuesforming disulphide bonds, have been termed '4-disulphide core' proteins. Protease inhibition occurs by insertion of the inhibitory loop into the active site pocket and interference with the catalytic residues of the protease.

                         10        20        30
                 ....*....|....*....|....*....|....*....
gi 755499091  90 CPRVRVKCEVEERNECTRHRQCPNKKRCCLFSCGKKCMD 128
Cdd:cd00199   20 CPMVNPPSLGIPPNRCSSDSDCPGDKKCCENGCGKSCLT 58

Kunitz domain of epididymal protease inhibitor eppin and similar proteins; This subfamily includes the Kunitz inhibitor domain protein eppin (also called Cancer/testis antigen 71 or CT71, epididymal protease inhibitor, protease inhibitor WAP7, serine protease inhibitor-like with Kunitz and WAP domains 1, or WAP four-disulfide core domain protein 7) as well as WAP four-disulfide core domain proteins 6A and 6B in mice, and similar proteins. Eppin is a serine protease inhibitor that plays an essential role in male reproduction and fertility. It modulates the hydrolysis of seminal fluid protein semenogelin 1 (SEMG1) by the serine protease kallikrein-related peptidase 3 (KLK3, PSA), provides antimicrobial protection for spermatozoa in the ejaculate coagulum, and binds SEMG1, thereby inhibiting sperm motility. Thus, eppin could potentially be used as a target for male contraception. These domains are similar to Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor) that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|....*..
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCKKKQ 188
Cdd:cd22611    1 DVCSLPKESGPCMAYFPRWWYDKETNTCSKFIYGGCQGNNNNFQSEAICQNICKKKR 57

Kunitz/Bovine pancreatic trypsin inhibitor (BPTI) domain; This family contains the Kunitz domain which is a common structural fold found in a family of reversible serine protease inhibitors. This domain is thought to have evolved over 500 million years and is ubiquitous in all kingdoms of life and has been incorporated into many different genes. In general, each domain is encoded by a single exon. Some genes encode proteins with a single Kunitz domain, e.g. bovine pancreatic trypsin inhibitor (BPTI), trophoblast Kunitz domain protein (TKDP), amyloid beta-protein precursor (ABPP), as well as Kunitz-type venom peptides such as dendrotoxin. Genes that encode multiple Kunitz domains include hepatocyte growth factor activator inhibitors HAI1 and HAI2 (two domains), tissue factor pathway inhibitor TFPI1 and TFPI2 (three domains) and Caenorhabditis elegans papilin (eleven domains). In addition, the Kunitz domain has been integrated into multi-domain proteins, e.g. the collagen alpha3(VI), alpha1(VII) and alpha1(XXVIII) chains, WFIKKN1 (containing WAP, Follistatin/Kazal, Immunoglobulin, two Kunitz and NTR domains) and papilin. Furthermore, each domain within a multi-Kunitz domain protein may exhibit different protease activity, such as for the three tandemly repeated domains within both tissue factor pathway inhibitors 1 and 2. The Kunitz domain is a representative of alpha/beta proteins with irregular secondary structure stabilized by three disulfide bonds and presenting three peptide loops that can be varied without introducing much destabilization to the scaffold. Protease inhibitors meet the scaffold criteria in that they are small, stable and capable of evolving the binding activity of exposed peptide loops through targeted randomization to construct combinatorial libraries. Kunitz domain-based scaffolds have been successfully utilized to construct and select a library of protease inhibitors with the potential for therapeutic application.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|.
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd00109    1 CLLPPDPGPCRAYFPRWYYNSETGQCEEFIYGGCGGNANNFETKEECEATC 51

first Kunitz domain of Carboxypeptidase Inhibitor SmCI and similar domains; This group includes Sabellastarte magnifica carboxypeptidase inhibitor (SmCI), a tri-domain BPTI-Kunitz inhibitor capable of inhibiting serine proteases and A-like metallocarboxypeptidases. While the BPTI-Kunitz family of proteins includes voltage gated channel blockers and inhibitors of serine proteases, SmCI is the only BPTI-Kunitz protein capable of inhibiting metallocarboxypeptidases. Binding studies show that SmCI is able to bind three trypsin molecules under saturating conditions, but only one elastase interacts with the inhibitor. Additionally, SmCI can bind serine proteases and carboxypeptidases at the same time (at least in the ratio 1:1:1), thus becoming the first protease inhibitor that simultaneously blocks these two mechanistic classes of enzymes. This model contains the first Kunitz domain of SmCI, which has a structure similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|...
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22601    2 DVCDLPADRGPCTAYIPRWFYNKTTKKCEKFVYGGCQGNKNRFETKDDCLANC 54

first Kunitz domain of Rhipicephalus microplus boophilin and similar proteins; This group includes venom serine protease inhibitors such as Rhipicephalus microplus and Ixodes scapularis boofilin, among others. Boophilin prevents blood clot formation to allow successful feeding and digestion through its inhibition activity of thrombin and other host anticoagulating factors like kallikrein, coagulation factor VII, or plasmin; it interacts with the host thrombin and trypsin. The structures of these domains are similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds. Rhipicephalus microplus boophilin contains two Kunitz domains; this model represents the first repeat.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|....*.
gi 755499091 130 RQDVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCK 185
Cdd:cd22599    2 RNGICRLPADEGICRALIPRFYFNTETGQCTEFIYGGCGGNENNFETIEECEKACG 57

Kunitz-type domain 1 (KD1) of hepatocyte growth factor activator inhibitor type 2 (HAI-2), and similar proteins; This model includes the Kunitz domain 1 (KD1) of hepatocyte growth factor activator inhibitor type 2 (HAI-2 or HAI2, also known as placental bikunin or Kunitz-type protease inhibitor 2). HAI-2 is composed of two Kunitz domains that strongly inhibit many serine proteases with sub-nanomolar affinities. HAI-2 Kunitz domain 1 (KD1) has been found to be the domain responsible for inhibition of hepatocyte growth factor (HGF) activator; activated HGF/scatter factor (HGF/SF) binds to its receptor tyrosine kinase MET to induce dimerization and initiate phosphorylation cascades leading to comprehensive cellular changes that, in the deregulated context of cancer, drive malignant transformation and progression. HAI-2 has been found to be a natural tumor suppressor in renal cell carcinoma, breast cancer and prostate cancer; its loss leads to tumor growth and progression in part due to increased MET signaling. HAI-2 is also a specific substrate for mesotrypsin, which is up-regulated with progression in prostate cancers and shown to contribute to invasion and metastasis; these activities of mesotrypsin may in part be mediated through cleavage and inactivation of HAI-2, resulting in increases in HGF/SF activation and MET signaling. HAI-2 is a physiological inhibitor of hepsin and matriptase, two type II transmembrane serine proteases that, like HGF activator, can convert latent pro-HGF/SF into the two-chain active signaling heterodimer. The structures of these KD1 domains are similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|...
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22621    1 DFCHLPKVVGRCRASFPRWWYNATSQSCQEFIFGGCKGNLNNFLSEQECLQKC 53

first Kunitz domain of bikunin and similar proteins; This subfamily includes the N-terminal domain of bikunin (also known as inter-alpha-trypsin inhibitor light chain (ITI-LC) or urinary trypsin inhibitor), a plasma protease inhibitor, that is associated with inflammation and stabilizes the extracellular matrix. It is encoded together with alpha-1-microglobulin (A1M) by an alpha-1-microglobulin/bikunin precursor (AMBP) gene that is tightly controlled by several hepatocyte-enriched nuclear (HEN) factors, and cleaved by a furin-like protease that releases the two mature molecules. Bikunin is a Kunitz-type serine protease inhibitor, found in vertebrate serum and urine, modified by a chondroitin sulfate (CS) chain. The structures of these toxins are similar to that of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds. Bikunin contains two Kunitz domains; this model represents the first repeat.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|....
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCK 185
Cdd:cd22596    1 DSCKLPPDAGPCFGMIQRYFYNSSSMACQTFNYGGCLGNQNNFVTEKECLQTCR 54

second Kunitz domain of WAP, Kazal, immunoglobulin, Kunitz and NTR domain-containing proteins; This subfamily includes WAP, Kazal, immunoglobulin, Kunitz and NTR domain-containing protein 1 (WFIKKN1, WFKN1), WFIKKN2 (WFKN2), and similar proteins. WFIKKN proteins are protease inhibitors that contain two distinct Kunitz-type protease inhibitor domains. They may have serine protease- and metalloprotease-inhibitor activity. This model represents the second Kunitz (KU2) domain, which has been shown to inhibit trypsin, but not chymotrypsin, elastase, plasmin, pancreatic kallikrein, lung tryptase, plasma kallikrein, thrombin, urokinase or tissue plasminogen activator. However, the inhibition constant of this domain for bovine trypsin is about five orders of magnitudes lower than that of bovine pancreatic trypsin inhibitor (BPTI) for trypsin. This could be due to unfavorable side-chain conformation of a tryptophan at P2' site which is incompatible with a trypsin complex; typical trypsin inhibitors of the Kunitz family feature a tyrosine residue or other less bulky residues at this site. The structure of KU2 is similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|..
gi 755499091 133 VCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22606    1 ICSLPAVQGPCKAWEPRWAYNSLLKQCQSFVYGGCEGNENNFESKEACEDAC 52

Kunitz-type actitoxins such as Anemonia viridis U-actitoxin-Avd3l, and similar proteins; This model includes the Kunitz-type actitoxins such as Anemonia viridis U-actitoxin-Avd3l (also called U-AITX-Avd3l or AsKC9), Anthopleura elegantissima KappaPI-actitoxin-Ael3a (also called KappaPI-AITX-Ael3a or Kunitz-type serine protease inhibitor APEKTx1) and Anthopleura aff. xanthogrammica PI-actitoxin-Axm2b (also called PI-AITX-Axm2b or Kunitz-type proteinase inhibitor AXPI-II). U-AITX-Avd3l and KappaPI-AITX-Ael3a are dual-function toxins that inhibit both the serine protease trypsin and voltage-gated potassium channels Kv1.2/KCNA2. These proteins are similar to Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor) that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|..
gi 755499091 133 VCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22633    4 ICLLPKDVGGCRARFPRYYYNSSTRRCEKFRYGGCGGNANNFHTLEECEKVC 55

Caenorhabditis elegans Kunitz domain 11 of papilin (also called abnormal cell migration protein 6 or mig-6), Amblyomma hebraeum amblin domain 1, and similar proteins; This model includes Caenorhabditis elegans Kunitz domain 11 of papilin (also called abnormal cell migration protein 6 or mig-6) and domain 1 of Amblyomma hebraeum amblin, and similar proteins. C. elegans papilin (also called abnormal cell migration protein 6) mig-6 encodes long (MIG-6L) and short (MIG-6S) isoforms of the extracellular matrix protein papilin, each required for distinct aspects of distal tip cell (DTC) migration and both isoforms have an N-terminal papilin cassette, lagrin repeats and six C-terminal Kunitz-type serine proteinase inhibitory domains. It plays a role in embryogenesis, the second phase of distal cell tip migration and is required for distribution of the metalloproteinase, mig-17, during organogenesis. Amblin contains two Kunitz-like domains and specifically inhibits thrombin. These domains are similar to Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|.
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22638    1 CTLKPETGPCRAYIEKWYYDPSTQSCKTFIYGGCGGNGNRFDSEEDCQETC 51

Kunitz domain of papilin, and similar proteins; This model includes the Kunitz domain found in human and mouse papilin, and similar proteins. Papilin is an extracellular matrix glycoprotein that has been found in many organisms to be involved in thin matrix layers during gastrulation, matrix associated with wandering, phagocytic hemocytes, basement membranes and space-filling matrix during Drosophila development. It is a multidomain protein that primarily occurs in basement membranes. Papilins interact with several extracellular matrix components and ADAMTS enzymes, influences cell rearrangements and may modulate metalloproteinases during organogenesis. Papilins exist in mammals and invertebrates as a set of related, though not necessarily identical proteins. Mammalian papilin contains a single Kunitz domain, while other papilins such as that from Caenorhabditis elegans, contains multiple Kunitz domains. These domains are similar to Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor) that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

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                 ....*....|....*....|....*....|....*....|....*....|..
gi 755499091 134 CSLPQDPG-PCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22635    1 CLLDKDAGtVCGDYVQRWYYDPATGACNRFWYGGCGGNANRFATEAECLRTC 52

Kunitz-type domain from the alpha3 chain of human type VI collagen, and similar proteins; This model includes the Kunitz-type domain from the alpha3 chain of type VI collagen (collagen alpha 3(VI) chain), encoded by COL6A3 gene. Collagen VI is a widely expressed member of the triple helix-containing protein superfamily of collagens and forms beaded microfibrils that anchor large interstitial structures. Immediately after fibril formation, the Kunitz domain can be cleaved off. Mutations in the alpha1, alpha2, and alpha3 chains of collagen VI cause myopathies ranging from the severe Ullrich congenital muscular dystrophy to the milder Bethlem myopathy, including intermediate forms. Early onset isolated dystonia, a neurological disease, has been shown to be caused by mutations in the alpha3 chain. Findings also indicated potential associations between COL6A3 polymorphisms and lung cancer risk. This domain is similar to that of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|...
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22629    1 DICKLPKDEGTCRDFVLKWYYDPETKSCARFWYGGCGGNENRFDSQEECEKVC 53

Kunitz domain 1 of hepatocyte growth factor activator inhibitor-1 (HAI-1); This model includes Kunitz domain 1 (KD1) of hepatocyte growth factor activator inhibitor type 1 (HAI1 or HAI-1, also known as Kunitz-type protease inhibitor 1), a membrane-bound multidomain protein essential to the integrity of the basement membrane during placental development. HAI-1 contains an extracellular region and several internal domains that include two Kunitz domains separated in sequence but spatially closed to each other, and their interdomain interactions have evolved to stimulate the inhibitory activity of an integrated Kunitz. KD1, the major inhibitory domain of HAI-1, is involved in auto-inhibition of the extracellular region via steric blockage of its active site in the HAI-1 compact tertiary structure; presence of the target protease causes changes in the HAI-1 structure to an extended conformation. HAI-1 has been shown to inhibit several serine proteases such as matripase, hepsin, trypsin, hepatocyte growth factor activator (HGFA), and prostasin. It is also important in maintaining postnatal homeostasis in many tissues, including keratinization of the epidermis, hair development, colonic epithelium integrity, proliferation and cell fate of neural progenitor cells, and tissue injury and repair. The interaction between HAI-1 and matriptase is critical for tissue morphogenesis and cellular biology. HAI-1:matriptase ratio imbalance results in tumorigenesis; slight overexpression of matriptase relative to HAI-1 causes spontaneous squamous cell carcinoma, a phenotype that can be effectively reversed back to wild type by additional expression of HAI-1, indicating the need for a tight functional relationship between the two to maintain homeostasis. The structures of these domains are similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|..
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCK 185
Cdd:cd22623    6 CLAPKKVGPCRGSFPRWHYNAASGKCEEFVFGGCKGNKNNYLSEEECLSACR 57

Stichodactyla helianthus Kunitz inhibitor protein ShPI-1, Heteractis crispa protease inhibitor stichotoxin-Hcr2e, and similar proteins; This model includes Kunitz inhibitor protein ShPI-1, the major protease inhibitor from the sea anemone Stichodactyla helianthus, as well as protease inhibitor stichotoxin-Hcr2e (also called PI- stichotoxin-Hcr2e, PI-SHTX-Hcr2e, or Kunitz-type serine protease inhibitor InhVJ) and HCRG1 from Heteractis crispa. ShPI-1 has an unusually broad specificity toward several serine proteases, including trypsin, chymotrypsin, human neutrophil elastase, kallikrein and plasmin, and can also bind aspartic and cysteine proteases, such as pepsin and papain, respectively. PI-SHTX-Hcr2e and HCRG1 inhibit trypsin and chymotrypsin, but do not inhibit the serine proteases plasmin, thrombin, kallikrein, the cysteine proteinase papain, and the aspartic protease pepsin. The structures of these domains are similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|...
gi 755499091 133 VCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCK 185
Cdd:cd22618    1 ICSEPKVVGPCKAYFPRFYFDSETGKCTPFIYGGCGGNGNNFETLHACRAICR 53

Kunitz domain 2 of hepatocyte growth factor activator inhibitor-1 (HAI1); This model includes Kunitz domain 2 (KD2) of hepatocyte growth factor activator inhibitor type 1 (HAI-1 or HAI1, also known as Kunitz-type protease inhibitor 1), a membrane-bound multidomain protein essential to the integrity of the basement membrane during placental development. HAI-1 contains an extracellular region and several internal domains that include two Kunitz domains separated in sequence but spatially closed to each other, and their interdomain interactions have evolved to stimulate the inhibitory activity of an integrated Kunitz. While the Kunitz domain 1 (KD1) is the major inhibitory domain of HAI-1 and involved in auto-inhibition of the extracellular region via steric blockage of its active site in the HAI-1 compact tertiary structure, studies show that deletion of HAI-1 Kunitz domain 2 (KD2) and the extracellular region enhanced inhibition of matriptase. HAI-1 KD2 has been shown to have potent inhibitory activity against trypsin, but it cannot inhibit hepatocyte growth factor activator (HGFA), and matriptase. HAI-1 is also important in maintaining postnatal homeostasis in many tissues, including keratinization of the epidermis, hair development, colonic epithelium integrity, proliferation and cell fate of neural progenitor cells, and tissue injury and repair. The interaction between HAI-1 and matriptase is critical for tissue morphogenesis and cellular biology. HAI-1:matriptase ratio imbalance results in tumorigenesis; slight overexpression of matriptase relative to HAI-1 causes spontaneous squamous cell carcinoma, a phenotype that can be effectively reversed back to wild type by additional expression of HAI-1, indicating the need for a tight functional relationship between the two to maintain homeostasis. The structure of KD2 is similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|..
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCK 185
Cdd:cd22624    2 CTEPPVTGPCRASFTRWYYDPLSRKCHRFTYGGCDGNENNFETEDECMETCS 53

Kunitz-type domain from the alpha6 chain of human type VI collagen, and similar proteins; This model includes the Kunitz-type domain from the alpha6 chain of type VI collagen (collagen alpha 6(VI) chain), encoded by COL6A6 gene, and similar proteins. Collagen VI is a widely expressed member of the triple helix-containing protein superfamily of collagens and forms beaded microfibrils that anchor large interstitial structures. Immediately after fibril formation, the Kunitz domain can be cleaved off. This domain is similar to that of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor) that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|....*
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVCKK 186
Cdd:cd22630    1 DACSLDQDEGECQNYVLKWYYDQEQKECSQFWYGGCGGNKNRFETQEECEALCVK 55

Kunitz domain 2 (KD2) of tissue factor pathway inhibitor 2 (TFPI2) and similar proteins; This model represents the Kunitz-type domain 2 (KD2) of tissue factor pathway inhibitor 2 (TFPI2 or TFPI-2) and similar proteins. TFPI2 exhibits inhibitory activity primarily toward trypsin, plasmin, and factor VIIa (FVIIa)/tissue factor (TF) via its KD1. It is believed to be the major inhibitor of plasmin in the extracellular matrix (ECM) but has little inhibitory activity toward urokinase-type plasminogen activator, tissue-type plasminogen activator, or thrombin. While TFPI2 specifically inhibits the proteases via the P1 arginine residue in KD1, domains KD2 and KD3 appear to have no discernible inhibitory activity and may serve to bind to nearby proteins to localize TFPI2 in the ECM. This domain is similar to that of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor) that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

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                 ....*....|....*....|....*....|....*....|....*....|....
gi 755499091 131 QDVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22617    1 PKVCREVPDEGPCRALITRYFYNMTSMRCEEFTYGGCYGNGNNFRDKSSCISAC 54

Kunitz-type domain from the alpha1 chain of type VII collagen, and similar proteins; This model includes the Kunitz-type domain from the alpha1 chain of type VII collagen (collagen alpha-1(VII) chain also called long-chain collagen or LC collagen) and similar proteins. LC collagen, encoded by the COL7A1 gene, is a stratified squamous epithelial basement membrane protein that forms anchoring fibrils which may contribute to epithelial basement membrane organization and adherence by interacting with extracellular matrix (ECM) proteins such as type IV collagen. So far, over 800 COL7A1 mutations have been reported, including missense, nonsense, splicing, insertion, and deletion mutations which to varying degrees leads to deficiency of type VII collagen. Epidermolysis bullosa acquisita (EBA) is an autoimmune acquired blistering skin disease resulting from autoantibodies to type VII collagen. The COL7A1 protein contains a Kunitz domain, the deactivation of which induces tumorigenesis. This domain is similar to that of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|...
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22627    1 DPCLLPMDEGSCSDYTLLWYYHQKAGECRPFVYGGCGGNANRFSSKEDCELRC 53

second Kunitz domain of bikunin and similar proteins; This subfamily includes the C-terminal domain of bikunin (also known as inter-alpha-trypsin inhibitor light chain (ITI-LC) or urinary trypsin inhibitor), a plasma protease inhibitor, that is associated with inflammation and stabilizes the extracellular matrix. Bikunin is encoded together with alpha-1-microglobulin (A1M) by an alpha-1-microglobulin/bikunin precursor (AMBP) gene that is tightly controlled by several hepatocyte-enriched nuclear (HEN) factors, and cleaved by a furin-like protease that releases the two mature molecules. Bikunin is a Kunitz-type serine protease inhibitor, found in vertebrate serum and urine, modified by a chondroitin sulfate (CS) chain. The structures of these toxins are similar to that of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds. Bikunin contains two Kunitz domains; this model represents the second repeat.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|...
gi 755499091 132 DVCSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22597    2 AACRLPIVPGPCKGFVDLWAFDAVQGKCVPFSYGGCQGNGNKFYSEKECEEYC 54

second Kunitz domain of Carboxypeptidase Inhibitor SmCI and similar domains; This group includes Sabellastarte magnifica carboxypeptidase inhibitor (SmCI), a tri-domain BPTI-Kunitz inhibitor capable of inhibiting serine proteases and A-like metallocarboxypeptidases. While the BPTI-Kunitz family of proteins includes voltage gated channel blockers and inhibitors of serine proteases, SmCI is the only BPTI-Kunitz protein capable of inhibiting metallocarboxypeptidases. Binding studies show that SmCI is able to bind three trypsin molecules under saturating conditions, but only one elastase interacts with the inhibitor. Additionally, SmCI can bind serine proteases and carboxypeptidases at the same time (at least in the ratio 1:1:1), thus becoming the first protease inhibitor that simultaneously blocks these two mechanistic classes of enzymes. This model contains the second Kunitz domain of SmCI, which has a structure similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|.
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22602    1 CSLPSKVGPCRVSARRWFHNPETEKCEVFIYGGCHGNANRFATETECQEVC 51

trophoblast Kunitz domain protein (TKDP) and similar proteins; This model contains the trophoblast Kunitz domain protein 1 (TKDP-1) and splice variant TKDP-4, among others, which are Kunitz inhibitor domain proteins. TKDP-1 is expressed in the trophectoderm which forms the outer epithelial layer of the trophoblast, and may play a role in mediating maternal-conceptus interactions in the immediate preimplantation period. However, it does not appear to have proteinase inhibitory activity. These domains are similar to those of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor) that shows an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|.
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22609    2 CLEPKVVGVCKASMTRYFYNAQTGHCEQFVYGGCGGNRNNFLTLEDCMKTC 52

Kunitz protease inhibitor (KPI) domain 3 (KPI-3 or K3) of tissue factor pathway inhibitor (TFPI) and TFPI2, and similar proteins; This model represents the third Kunitz-type domain (K3 or KPI-3) of tissue factor pathway inhibitor (TFPI or TFPI1), also known as extrinsic pathway inhibitor (EPI) or lipoprotein-associated coagulation inhibitor (LACI), and of TFPI2 (or TFPI-2). TFPI1 down-regulates the extrinsic coagulation pathway via inhibition of activated factor X (FXa or Xa) and FVIIa (VIIa). It inhibits activated FXa via a "slow-tight binding mechanism", i.e. rapid formation of a loose FXa-TFPI1 complex that then slowly isomerizes to a tight FXa-TFPI1* complex. Subsequent inhibition of FVIIa is facilitated by the presence of tissue factor (TF) and FXa, which together rapidly and efficiently form a quaternary FXa-TFPI1-TF-FVIIa complex in which the activity of FXa and FVIIa are inhibited. TFPI1 consists of 3 Kunitz-type protease inhibitor (KPI) domains in a tandem arrangement; while the K1 domain of TFPI has been shown to bind and inhibit FVIIa and the K2 domain similarly inhibits FXa, the K3 domain has no known inhibitory function. However, Protein S, which functions as a cofactor for TFPI to efficiently enhance TFPI inhibition of FXa and FXa activated TF-VIIa, is dependent on direct interactions with two important residues within K3, a Glutamate and an Arginine. This model also includes TFPI2 Kunitz domain 3 (KD3). TFPI2 exhibits inhibitory activity primarily toward trypsin, plasmin, and factor VIIa (FVIIa)/tissue factor (TF) via its KD1. It is believed to be the major inhibitor of plasmin in the extracellular matrix (ECM) but has little inhibitory activity toward urokinase-type plasminogen activator, tissue-type plasminogen activator, or thrombin. While TFPI2 specifically inhibits the proteases via the P1 arginine residue in KD1, domains KD2 and KD3 appear to have no discernible inhibitory activity and may serve to bind to nearby proteins to localize TFPI2 in the ECM. The structure of this domain is similar to that of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|.
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22615    4 CLSPKDEGLCSASVTRYYYNSATKTCEPFNYTGCGGNNNNFTSKKDCLRVC 54

bovine pancreatic trypsin inhibitor; This model contains bovine pancreatic trypsin inhibitor (BPTI, also known as pancreatic Kunitz inhibitor, aprotinin, or trypsin-kallikrein inhibitor), a small protein that inhibits the action of the trypsin, and is thus a member of the serine protease family of inhibitors. This class of enzymes contains conserved cysteine residues that form 3 disulfide bonds to stabilize the three-dimensional structure. BPTI has a relatively broad specificity, inhibiting trypsin as well as chymotrypsin, and elastase-like serine (pro)enzymes capable of very different primary specificity. It reacts rapidly with serine proteases to form stable complexes, but the enzyme:inhibitor complex formation may involve several intermediates corresponding to discrete reaction steps. Furthermore, BPTI inhibits the nitric oxide synthase type-I and -II action, and impairs K+ transport by Ca2+-activated K+ channels. Clinically, BPTI is used in certain surgical interventions, such as cardiopulmonary surgery and orthotopic liver transplantation since it significantly reduces hemorrhagic complications.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|.
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22592    2 CLEPPYTGPCKARIIRYFYNAKSGLCETFVYGGCRAKRNNFLSAEDCMRTC 52

dendrotoxins I, K, B and similar proteins; This group includes toxins isolated from snake venoms, such as dendrotoxins (DTXs) I, K and B, mambaquaretin-1 (MQ-1) and calcicludine. The dendrotoxins have little or no anti-protease activity but have been shown to block certain subtypes of voltage dependent potassium channels in neurons. Dendroaspis angusticeps (green mamba) alpha-dendrotoxin is a neurotoxin that enhances acetylcholine release at neuromuscular junctions. Studies with cloned K(+) channels show that this toxin blocks Kv1.1, Kv1.2 and Kv1.6 channels in the nanomolar range, whereas Dendroaspis polylepis (black mamba) dendrotoxin K preferentially blocks Kv1.1 channels. Also, structural analogs of dendrotoxins have facilitated defining the molecular recognition properties of different types of K(+) channels, and therefore, dendrotoxins are widely used as probes for studying the function of K(+) channels in physiology and pathophysiology. The structures of these toxins are similar to that of Kunitz-type proteinase inhibitors such as BPTI (bovine pancreatic trypsin inhibitor), showing an alpha/beta fold with irregular secondary structure stabilized by three disulfide bonds.

                         10        20        30        40        50
                 ....*....|....*....|....*....|....*....|....*....|.
gi 755499091 134 CSLPQDPGPCLAYLPRWWYNQETDLCTEFIYGGCQGNPNNFPSEGICTVVC 184
Cdd:cd22595    4 CKLPVRPGPCKAFISAFYYNWKAKKCHPFTYSGCGGNANRFKTIEECRRTC 54

WAP-type (Whey Acidic Protein) 'four-disulfide core'; WAP belongs to the group of Elafin or elastase-specific inhibitors.

                          10        20        30        40
                  ....*....|....*....|....*....|....*....|
gi 755499091   89 TCPRVRVKCEVeeRNECTRHRQCPNKKRCCLFSCGKKCMD 128
Cdd:pfam00095   4 CCPRLGARGCC--RSCCSSDDDCPGRQKCCSNGCGSVCVP 41

whey acidic protein-type four-disulfide core domains. Members of the family include whey acidic protein, elafin (elastase-specific inhibitor), caltrin-like protein (a calcium transport inhibitor) and other extracellular proteinase inhibitors. A group of proteins containing 8 characteristically-spaced cysteine residuesforming disulphide bonds, have been termed '4-disulphide core' proteins. Protease inhibition occurs by insertion of the inhibitory loop into the active site pocket and interference with the catalytic residues of the protease.

                         10        20        30
                 ....*....|....*....|....*....|....*....
gi 755499091  90 CPRVRVKCEVEERNECTRHRQCPNKKRCCLFSCGKKCMD 128
Cdd:cd00199   20 CPMVNPPSLGIPPNRCSSDSDCPGDKKCCENGCGKSCLT 58

Four-disulfide core domains;

                           10        20        30
                   ....*....|....*....|....*....|
gi 755499091    99 VEERNECTRHRQCPNKKRCCLFSCGKKCMD 128
Cdd:smart00217  16 GNPPNKCSSDSQCPGVKKCCFNGCGKSCLT 45