ADAMTS-13 ja COVID-19. PubMed haku

Microvascular dysfunction in COVID-19: the MYSTIC study.

Rovas A, Osiaevi I, Buscher K, Sackarnd J, Tepasse PR, Fobker M, Kühn J, Braune S, Göbel U, Thölking G, Gröschel A, Pavenstädt H, Vink H, Kümpers P. Angiogenesis. 2021 Feb;24(1):145-157. doi: 10.1007/s10456-020-09753-7. Epub 2020 Oct 14. PMID: 33058027 Free PMC article.

METHODS: Hospitalized adult patients with moderate-to-severe or critical COVID-19 (n = 23) were enrolled non-consecutively in this prospective, observational, cross-sectional, multi-center study. ...CONCLUSIONS: Our data clearly show severe alterations …

2

Prognostic value of von Willebrand factor and ADAMTS13 in patients with COVID-19: A systematic review and meta-analysis.

Xu X, Feng Y, Jia Y, Zhang X, Li L, Bai X, Jiao L. Thromb Res. 2022 Oct;218:83-98. doi: 10.1016/j.thromres.2022.08.017. Epub 2022 Aug 18. PMID: 36027630 Free PMC article. Review.

The estimated pooled means indicated increased plasma levels of VWF:Ag, VWF:Rco, and VWF:Ag/ADAMTS13:Ac ratio, and decreased plasma levels of ADAMTS13:Ac in COVID-19 patients with unfavorable outcomes when compared to those with favorable outcomes (com …

Background: Endotheliopathy and coagulopathy appear to be the main causes for critical illness and death in patients with coronavirus disease 2019 (COVID-19). The adhesive ligand von Willebrand factor (VWF) has been involved in immunothrombosis responding to endothelial injury. Here, we reviewed the current literature and performed meta-analyses on the relationship between both VWF and its cleaving protease ADAMTS13 (a disintegrin and metalloproteinase with thrombospondin type 1 motif, member 13) with the prognosis of COVID-19. Methods: We searched MEDLINE, Cochrane Library, Web of Science, and EMBASE databases from inception to 4 March 2022 for studies analyzing the relationship between VWF-related variables and composite clinical outcomes of patients with COVID-19. The VWF-related variables analyzed included VWF antigen (VWF:Ag), VWF ristocetin cofactor (VWF:Rco), ADAMTS13 activity (ADAMTS13:Ac), the ratio of VWF:Ag to ADAMTS13:Ac, and coagulation factor VIII (FVIII). The unfavorable outcomes were defined as mortality, intensive care unit (ICU) admission, and severe disease course. We used random or fixed effects models to create summary estimates of risk. Risk of bias was assessed based on the principle of the Newcastle-Ottawa Scale. Results: A total of 3764 patients from 40 studies were included. The estimated pooled means indicated increased plasma levels of VWF:Ag, VWF:Rco, and VWF:Ag/ADAMTS13:Ac ratio, and decreased plasma levels of ADAMTS13:Ac in COVID-19 patients with unfavorable outcomes when compared to those with favorable outcomes (composite outcomes or subgroup analyses of non-survivor versus survivor, ICU versus non-ICU, and severe versus non-severe). In addition, FVIII were higher in COVID-19 patients with unfavorable outcomes. Subgroup analyses indicated that FVIII was higher in patients admitting to ICU, while there was no significant difference between non-survivors and survivors. Conclusions: The imbalance of the VWF-ADAMTS13 axis (massive quantitative and qualitative increases of VWF with relative deficiency of ADAMTS13) is associated with poor prognosis of patients with COVID-19.

3

Increased VWF and Decreased ADAMTS-13 in COVID-19: Creating a Milieu for (Micro)Thrombosis.

Favaloro EJ, Henry BM, Lippi G. Semin Thromb Hemost. 2021 Jun;47(4):400-418. doi: 10.1055/s-0041-1727282. Epub 2021 Apr 23. PMID: 33893632 Review.

However, ADAMTS-13 deficiency may result from other pathological processes. Of relevance is the recent finding that COVID-19 (coronavirus disease 2019) is associated with both increased levels and activity of VWF as well as generally decreased  (or occasionally normal) activity levels of ADAMTS-13. Thus, in COVID-19 there is an alteration in the VWF/ADAMTS-13 axis, most often described by increased VWF/ADAMTS-13 ratio (or reduced ADAMTS-13/VWF ratio). COVID-19 is also associated with high prothrombotic risk. Thus, the imbalance of VWF and ADAMTS-13 in COVID-19 may be providing a milieu that promotes (micro)thrombosis, in a clinical picture resembling a secondary thrombotic microangiopathy in some patients. This review therefore assesses the literature on VWF, ADAMTS-13, and COVID-19. Whenever reported in COVID-19, VWF has always been identified as raised (compared with normal reference ranges or control populations). Reports have included VWF level (i.e., VWF antigen) and in some cases one or more VWF "activity" (e.g., collagen binding; platelet glycoprotein Ib [GPIb] binding, using ristocetin cofactor or more modern versions including VWF:GPIbR [recombinant] and VWF:GPIbM [mutant]). Whenever reported, ADAMTS-13 has been reported as "normal" or reduced; however, it should be recognized that "normal" levels may still identify a relative reduction in individual cases. Some reports also discuss the raised VWF/ADAMTS-13 (or reduced ADAMTS-13/VWF) ratio, but very few provide actual numerical data.

4

The ADAMTS13-von Willebrand factor axis in COVID-19 patients.

Mancini I, Baronciani L, Artoni A, Colpani P, Biganzoli M, Cozzi G, Novembrino C, Boscolo Anzoletti M, De Zan V, Pagliari MT, Gualtierotti R, Aliberti S, Panigada M, Grasselli G, Blasi F, Peyvandi F. J Thromb Haemost. 2021 Feb;19(2):513-521. doi: 10.1111/jth.15191. Epub 2020 Dec 18. PMID: 33230904 Free PMC article.

BACKGROUND: Severe coronavirus disease 2019 (COVID-19) is characterized by an increased risk of thromboembolic events, with evidence of microthrombosis in the lungs of deceased patients. ...CONCLUSIONS: We found a significant alteration of the V …

5

ADAMTS13 regulation of VWF multimer distribution in severe COVID-19.

Ward SE, Fogarty H, Karampini E, Lavin M, Schneppenheim S, Dittmer R, Morrin H, Glavey S, Ni Cheallaigh C, Bergin C, Martin-Loeches I, Mallon PW, Curley GF, Baker RI, Budde U, O'Sullivan JM, O'Donnell JS; Irish COVID-19 Vasculopathy Study (iCVS) investigators. J Thromb Haemost. 2021 Aug;19(8):1914-1921. doi: 10.1111/jth.15409. Epub 2021 Jun 20. PMID: 34053187 Free PMC article.

OBJECTIVES: This study investigated the hypothesis that ADAMTS-13 regulation of VWF multimer distribution may be impaired in severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection contributing to the o … We observed markedly increased VWF collagen-binding activity in patients with severe COVID-19 compared to controls (median 509.1 versus 94.3 IU/dl). Conversely, plasma ADAMTS-13 activity was significantly reduced (median 68.2 IU/dl). In keeping with an increase in VWF:ADAMTS-13 ratio, abnormalities in VWF multimer distribution were common in patients with COVID-19, with reductions in high molecular weight VWF multimers. Terminal sialylation regulates VWF susceptibility to proteolysis by ADAMTS-13 and other proteases. We observed that both N- and O-linked sialylation were altered in severe COVID-19. Furthermore, plasma levels of the ADAMTS-13 inhibitors interleukin-6, thrombospondin-1, and platelet factor 4 were significantly elevated.

6

Distinctive Biomarker Features in the Endotheliopathy of COVID-19 and Septic Syndromes.

Fernández S, Moreno-Castaño AB, Palomo M, Martinez-Sanchez J, Torramadé-Moix S, Téllez A, Ventosa H, Seguí F, Escolar G, Carreras E, Nicolás JM, Richardson E, García-Bernal D, Carlo-Stella C, Moraleda JM, Richardson PG, Díaz-Ricart M, Castro P. Shock. 2022 Jan 1;57(1):95-105. doi: 10.1097/SHK.0000000000001823. PMID: 34172614 Free PMC article.

Biomarkers distinguishing different COVID-19 phenotypes from sepsis syndrome remain poorly understood. ...METHODS: Patients with COVID-19 pneumonia (n = 49) were classified into moderate, severe, or critical (life-threatening) disease. Plasma samples were collected within 48 to 72 h of hospitalization to analyze endothelial activation markers, including soluble Vascular Cell Adhesion Molecule-1 (sVCAM-1), von Willebrand Factor (VWF), A disintegrin-like and metalloprotease with thrombospondin type 1 motif no. 13 (ADAMTS-13) activity, thrombomodulin (TM), and soluble TNF receptor I (sTNFRI); heparan sulfate (HS) for endothelial glycocalyx degradation; C5b9 deposits on endothelial cells in culture and soluble C5b9 for complement activation; circulating dsDNA for neutrophil extracellular traps (NETs) presence, and α2-antiplasmin and PAI-1 as parameters of fibrinolysis. We compared the level of each biomarker in all three COVID-19 groups and healthy donors as controls (n = 45). Results in critically ill COVID-19 patients were compared with other intensive care unit (ICU) patients with septic shock (SS, n = 14), sepsis (S, n = 7), and noninfectious systemic inflammatory response syndrome (NI-SIRS, n = 7). … All analyzed biomarkers were increased in COVID-19 patients versus controls (P < 0.001), except for ADAMTS-13 activity that was normal in both groups. The increased expression of sVCAM-1, VWF, sTNFRI, and HS was related to COVID-19 disease severity (P < 0.05). Several differences in these parameters were found between ICU groups: SS patients showed significantly higher levels of VWF, TM, sTNFRI, and NETS compared with critical COVID-19 patients and ADAMTS-13 activity was significantly lover in SS, S, and NI-SIRS versus critical COVID-19 (P < 0.001). Furthermore, α2-antiplasmin activity was higher in critical COVID-19 versus NI-SIRS (P < 0.01) and SS (P < 0.001), whereas PAI-1 levels were significantly lower in COVID-19 patients compared with NI-SIRS, S, and SS patients (P < 0.01). Conclusions: COVID-19 patients present with increased circulating endothelial stress products, complement activation, and fibrinolytic dysregulation, associated with disease severity. COVID-19 endotheliopathy differs from SS, in which endothelial damage is also a critical feature of pathobiology. These biomarkers could help to stratify the severity of COVID-19 disease and may also provide information to guide specific therapeutic strategies to mitigate endotheliopathy progression.

7

Low ADAMTS13 Activity Correlates with Increased Mortality in COVID-19 Patients.

Sweeney JM, Barouqa M, Krause GJ, Gonzalez-Lugo JD, Rahman S, Gil MR. TH Open. 2021 Mar 9;5(1):e89-e103. doi: 10.1055/s-0041-1723784. eCollection 2021 Jan. PMID: 33709050 Free PMC article.

The causes of coagulopathy associated with coronavirus disease 2019 (COVID-19) are poorly understood. We aimed to investigate the relationship between von Willebrand factor (VWF) biomarkers, intravascular hemolysis, coagulation, and organ damage in …

8

COVID-19 associated coagulopathy: Mechanisms and host-directed treatment.

Plášek J, Gumulec J, Máca J, Škarda J, Procházka V, Grézl T, Václavík J. Am J Med Sci. 2022 Jun;363(6):465-475. doi: 10.1016/j.amjms.2021.10.012. Epub 2021 Nov 6. PMID: 34752741 Free PMC article. Review.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is associated with specific coagulopathy that frequently occurs during the different phases of coronavirus disease 2019 (COVID-19 …

9

Impaired exercise capacity in post-COVID-19 syndrome: the role of VWF-ADAMTS13 axis.

Prasannan N, Heightman M, Hillman T, Wall E, Bell R, Kessler A, Neave L, Doyle A, Devaraj A, Singh D, Dehbi HM, Scully M. Blood Adv. 2022 Jul 12;6(13):4041-4048. doi: 10.1182/bloodadvances.2021006944. PMID: 35543533 Free PMC article.

Post-COVID syndrome (PCS), or long COVID, is an increasingly recognized complication of acute SARS-CoV-2 infection, characterized by persistent fatigue, reduced exercise tolerance, chest pain, shortness of breath, and cognitive slowing. Acute COVID-19 is strongly linked with an increased risk of thrombosis, which is a prothrombotic state quantified by an elevated von Willebrand factor (VWF) antigen (Ag)/ADAMTS13 ratio that is associated with severity of acute COVID-19 infection. We investigated whether patients with PCS also had evidence of a prothrombotic state associated with symptom severity. In a large cohort of patients referred to a dedicated post-COVID-19 clinic, thrombotic risk, including VWF(Ag)/ADAMTS13 ratio, was investigated. An elevated VWF(Ag)/ADAMTS13 ratio (≥1.5) was present in nearly one-third of the cohort and was 4 times more likely to be present in patients with impaired exercise capacity, as evidenced by desaturation ≥3% and/or an increase in lactate level >1 from baseline on a 1-minute sit-to-stand test and/or a 6-minute walk test (P < .0001). Of 276 patients, 56 (20%) had impaired exercise capacity, of which 55% (31/56) had a VWF(Ag)/ADAMTS13 ratio ≥1.5 (P < .0001). Factor VIII and VWF(Ag) were elevated in 26% and 18%, respectively, and support a hypercoagulable state in some patients with PCS. These findings suggest possible ongoing microvascular/endothelial dysfunction in the pathogenesis of PCS and suggest a role for antithrombotic therapy in the treatment of these patients.

10

ADAMTS13 factor deficiency in severe COVID-19 may not be immune mediated - report from a pilot study.

George A, Deepika C, Mohan G, Srishail R, Rajendran V, Shastry S, Balakrishnan JM, Rao S. Clin Hemorheol Microcirc. 2022;82(2):193-198. doi: 10.3233/CH-221514. PMID: 35754264

BACKGROUND: The assessment of ADAMTS13 factor activity and inhibitor levels was conducted in severe COVID-19 patients as an observational study. ..Results: A total of 14 patients were included and the average ADAMTS13 activity level at the time of admission was 28.54±30.74% (range 1.83-86.67%) which was reduced compared to controls (88.09±14.77). Nine patients had reduced ADAMTS13 factor activity (<40%) and 77.7% among them had severe deficiency (<10% activity). ADAMTS13 inhibitor was positive (>15 IU/mL) only in two patients and an overall mean value was 8.15±5.8. Elevated D-Dimer and length of hospital stay had significant correlation with ADAMTS13 activity (-0.247 and 0.306 respectively). No features of thrombotic microangiopathy were observed and hence no plasma exchange was performed. Conclusion: Reduced ADAMTS13 factor activity without inhibitor development may give a clue to the disease progress in COVID-19. …

ADAMTS-13 ja ADAMTS-13- vasta-aineet täsmädiagnostiikassa

https://www.researchgate.net/journal/Rambam-Maimonides-Medical-Journal-2076-9172 

ADAMTS-13 in the Diagnosis and Management of Thrombotic Microangiopathies

Article

Full-text available

• October 2014

• Galit Sarig

Thrombotic microangiopathies (TMAs) comprise a group of distinct disorders characterized by microangiopathic hemolytic anemia, thrombocytopenia, and microvascular thrombosis. For many years distinction between these TMAs, especially between thrombotic thrombocytopenic purpura (TTP) and hemolytic uremic syndrome (HUS), remained purely clinical and hard to make. Recent discoveries shed light on different pathogenesis of TTP and HUS. Ultra-large von Willebrand factor (UL-VWF) platelet thrombi, resulting from the deficiency of cleavage protease which is now known as ADAMTS-13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13), were found to cause TTP pathology, while Shiga toxins or abnormalities in regulation of the complement system cause microangiopathy and thrombosis in HUS. TMAs may appear in various conditions such as pregnancy, inflammation, malignancy, or exposure to drugs. These conditions might cause acquired TTP, HUS, or other TMAs, or might be a trigger in individuals with genetic predisposition to ADAMTS-13 or complement factor H deficiency. Differentiation between these TMAs is highly important for urgent initiation of appropriate therapy. Measurement of ADAMTS-13 activity and anti-ADAMTS-13 antibody levels may advance this differentiation resulting in accurate diagnosis. Additionally, assessment of ADAMTS-13 levels can be a tool for monitoring treatment efficacy and relapse risk, allowing consideration of therapy addition or change. In the past few years, great improvements in ADAMTS-13 assays have been made, and tests with increased sensitivity, specificity, reproducibility, and shorter turnaround time are now available. These new assays enable ADAMTS-13 measurement in routine clinical diagnostic laboratories, which may ultimately result in improvement of TMA management.

Zinkiinit , Metzinkiinit, Metalloproteaasit (MP)

 (Miten löysin tämän artikkelin tnään 23.11.2021: Olin etsimässä Sars-2 S-proteiinin  trimerisoitumisesta ensinnä arikkeleita ja siten  aloin pohtia  transmembraanisen  trimeroituneen tyven muodostumista ja etsiäö prokollageenin  trimerisoivaa domeenia; - samalla löysin  entsyymejä- , jotka pilkkovat tässä yhteydessä   mahdollista  trimeeriä koska se on transientti  fysiologisesti. Tosin sars-2  ei anna  trimeerien pilkoutua ainakaan ennen kuin uudet virionit ovat valmiita stabiilien trimeerien kanssa  ja toisaalta   infektoituminen  vanhalla trimeerillä on tehty.   Löysin siitä  fokuksesta  molekyylejä, jotka ovat  Sars-2 interaktioproteiineja ja samalla rokotevalmstuksessakin tarvittava. Yhdessä oli CUB-domeeni ja NTR-domeeni vapautui siitä   pilkkoutumalla. NTR-modulista sanottiin, että se oli " homologinen TIMP:n kanssa, mutta ei pystynyt  vaikuttamaan  metsinkiineihin".  Sen takia  etsin metsinkiinit esiin . Niin läysin tämän artikkelin). 

 

 

 https://www.jbc.org/article/S0021-9258(20)42509-8/fulltext

 

Minireviews| Volume 284, ISSUE 23, P15353-15357, June 2009

Catalytic Domain Architecture of Metzincin Metalloproteases

• F. Xavier Gomis-Rüth

Open AccessDOI:     https://doi.org/10.1074/jbc.R800069200

Metalloproteases cleave proteins and peptides, and deregulation of their function leads to pathology. An understanding of their structure and mechanisms of action is necessary to the development of strategies for their regulation. Among metallopeptidases are the metzincins, which are mostly multidomain proteins with ∼130–260-residue globular catalytic domains showing a common core architecture characterized by a long zinc-binding consensus motif, HEXXHXXGXX(H/D), and a methionine-containing Met-turn. Metzincins participate in unspecific protein degradation such as digestion of intake proteins and tissue development, maintenance, and remodeling, but they are also involved in highly specific cleavage events to activate or inactivate themselves or other (pro)enzymes and bioactive peptides.

 Metzincins are subdivided into families, and seven such families have been analyzed at the structural level: 

the astacins,

 ADAMs/adamalysins/reprolysins,

 serralysins,

 matrix metalloproteinases (MMPs) ,

 snapalysins, 

leishmanolysins, and

 pappalysins. 

These families are reviewed from a structural point of view.

Metalloproteases

Cleavage of peptide bonds is essential for life, and the factors responsible for peptide cleavage are ubiquitous. Among them are MPs, ² which are mostly zinc-dependent peptide-bond hydrolases. They participate in metabolism through both extensive and unspecific protein degradation and controlled hydrolysis of specific peptide bonds (1.).   Deregulation of such vast degrading potential leads to pathologies, and in addition, MPs may also act as virulence factors during poisoning and microbial infection. Such a wide range of biological functions makes structural studies of these proteins indispensable to any understanding of their function and to the design of novel, highly specific therapeutic agents to modulate their activity (2.).

Most MPs are members of a protease tribe, the zincins, and they possess a short consensus amino acidsequence, HEXXH. This motif contains two protein ligands of the catalytic zinc and a glutamate that acts as general base/acid during the catalytic process (3.,4.). A third metal ligand is a solvent molecule, further bound to and polarized by the glutamate. This solvent performs nucleophilic attack on the carbonyl carbon of the scissile bond of a bound substrate, leading to a tetrahedral gem-diolate reaction intermediate, which is stabilized by the positively charged metal ion and neighboring protein residues (5.). Subsequent evolution of the intermediate under assistance of the general base/acid eventually leads to bond disruption.

Zincins are divided into the

 gluzincin,

 aspzincin, and

 metzincin clans (4.,7.).

 The latter contains mostly multidomain proteins with an N-terminal prodomain engaged in latency maintenance, a catalytic protease domain, and farther downstream domains engaged in protein-protein and cell-cell interactions and other regulatory functions. The protease domain is characterized by a C-terminally extended zinc-binding motif, HEXXHXXGXX(H/D), with a hallmark glycine and a third zinc-binding histidine or aspartate. In addition, a methionine is present in a conserved downstream turn, the Met-turn (3.,7.,8.).

 Metzincins split into families, seven of which have been characterized at the structural level for at least one of their members: astacins, ADAMs/adamalysins/reprolysins, serralysins, matrix metalloproteinases, snapalysins, leishmanolysins, and pappalysins.

 In addition, a series of sequences reported from genome sequencing projects indicate there are further structurally yet uncharacterized families, tentatively referred to as

 fragilysins,

 gametolysins,

 archaemetzincins,

 thuringilysins, 

 coelilysins,

 ascomycolysins,

 helicolysins, and 

cholerilysins (for a detailed review, see Ref.7.).

The Metzincin Fold

To date, >200 structures of metzincins, comprising at least the catalytic domain, have been deposited with the Protein Data Bank (supplemental Table 1). In this review, seven lead structures of each of the aforementioned families are discussed: Astacus astacus astacin, Crotalus adamanteus adamalysin II, Pseudomonas aeruginosa aeruginolysin, human neutrophil collagenase (MMP-8), Streptomyces caespitosus neutral protease (snapalysin), Leishmania major leishmanolysin, and Methanosarcina acetivorans ulilysin (9.,10.,11.,12.,13.,14.,15.). These prototypes cover distinct kingdoms of life and represent archaea, bacteria, protozoa, crustaceans, reptiles, and mammals. The structures reveal that metzincins share a common scaffold and active-site environment, but each family has distinguishing structural elements. Mature catalytic domains are ∼130–260-residue globular moieties that bifurcate into an upper NSD and a lower CSD with respect to a central active-site cleft. Substrates bind horizontally to this cleft from left to right in an approximately extended conformation (supplemental Fig. 1 A). (All topological indications refer to the standard orientation displayed in this figure.)

 

The NSD displays a five-stranded twisted β-sheet at the top (except leishmanolysin, which has four strands) (supplemental Fig. 1A). All strands (βI–βV) except the fourth are parallel to each other and to any substrate that is bound in the cleft. The antiparallel strand, βIV, forms the lower edge of this subdomain and creates an upper rim or northern wall of the active-site crevice (16.). This strand binds a substrate in an antiparallel manner, mainly on its non-primed side. The loop segment connecting strands βIII and βIV (referred to as LβIIIβIV) leads to the appearance of bulge-like elements, which mainly affect subsites S₁′ and S₂′ (see Ref.17. for subsite nomenclature in proteases). This gives rise to extensive variations in enzyme-substrate interactions on the primed side of the active-site clefts. The NSD also contains two long α-helices, αA and αB, the backing helix and the active-site helix, respectively. Both are arranged on the concave side of the β-sheet in an identical manner in all metzincin structures (supplemental Fig. 1B). Helix αB superimposes well in all seven structures (Fig. 1A) and encompasses the first half of the zinc-binding motif, which includes the first two zinc-binding histidine residues (Fig. 1A and supplemental Fig. 1). At the end of helix αB, the polypeptide chain takes a sharp downward turn, mediated by the glycine of the consensus sequence. The main-chain angles of this residue in the different lead structures indicate that any other residue would be in a high-energy conformation and thus disfavored, with the exception of ulilysin (see below).

 

FIGURE 1Overlay of structural segments common to the seven prototypes as C^α plots. A, the active-site helix αB (including the side chains of the zinc-liganding histidines and aspartate and the general base/acid) and the Met-turn (with the methionine side chain). Blue, astacin; cyan, adamalysin II; red, leishmanolysin; green, MMP-8; white, aeruginolysin; yellow, snapalysin; orange, ulilysin. The magenta curved arrows indicate where an extra domain is inserted in leishmanolysin. B, the upper domain β-sheet (strands βI–βV, in cyan, blue, green, orange, and yellow, respectively), the posterior helix αA (red), and the C-terminal helix αC (purple) as found in the seven leads. Leishmanolysin lacks strand βII.

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The CSD starts after this glycine, and the chain leads to the third zinc ligand, a histidine or an aspartate, which approaches the metal from below (supplemental Fig. 1). This subdomain contains few repetitive secondary structure elements, mainly a C-terminal helix αC at the end of the polypeptide chain. Helices αB and αC are connected by structures that vary both in length and conformation. However, all structures coincide at a conserved 1,4-β-turn containing a methionine at position 3, the Met-turn, which is separated from the third zinc-binding histidine by connecting segments of 6–53 amino acids in the different structures. The Met-turn is superimposable (including the conformation of the methionine side chain) (Fig. 1A) and is positioned underneath the catalytic Zn²⁺, forming a hydrophobic pillow. However, no direct contact with the metal is observed. Mutation studies suggested a role for this methionine in the folding and stability of the catalytic domains, although the strict conservation of this residue remains to be explained (18.,19.). ³ The S₁′ pocket of metzincins is shaped at the top by a protruding bulge made by LβIIIβIV and at the bottom by a wall-forming segment made up of residues intercalated between the Met-turn and the C-terminal helix αC (16.). This segment diverges in structure and length (ranging from 11 to 37 residues between the Met-turn methionine and the first residue of helix αC) in all seven of the reference structures.

The Zinc-binding Site

The catalytic zinc ion lies roughly at the center of the bottom of the active-site cleft. It is coordinated by the N^ϵ2 atoms of the three consensus histidines (two histidine N^ϵ2 atoms and one aspartate O^δ2 atom in snapalysin) and the catalytic solvent molecule, substituted by other ligands in the enzyme/inhibitor-product complexes reported (supplemental Table 1). Some metzincins display an additional protein ligand at a slightly greater distance from the catalytic cation in the form of a tyrosine O^η atom, as seen in unbound astacin and serralysins and as hypothesized in ulilysin (21.). This tyrosine residue lies two positions ahead of the Met-turn methionine. In (unbound) snapalysin, a tyrosine lies two positions downstream of the metal-binding aspartate, though no longer within binding distance of the zinc ion. These tyrosine residues flip back and forth during substrate anchoring, cleavage, and product release in a motion referred to as the tyrosine switch. In this, they may play a role in substrate and catalytic solvent binding and stabilization of the tetrahedral intermediate or the product amino group (

7.,9.,22.,23.,24.). Such a role is performed by other, non-conserved residues in tyrosine-lacking metzincins.

Structural Relatedness among Families

A structure-based sequence alignment reveals that, with the exception of the region comprising the zinc-binding consensus sequence, sequence identity is negligible among metzincins despite evident structural relatedness. Based on this alignment, pairwise comparisons between leads (supplemental Fig. 2) show between 41 and 118 topologically equivalent C^α positions and root mean square deviations between 3.3 and 4.0Å, i.e. above the threshold at which two C^α positions can be considered as structurally equivalent (3.0 Å) (

25.). The sequence identities range between 3 and 20%, i.e. clearly below twilight values for sequence relatedness (25–35%) (26.). The closest structural pairs are aeruginolysin and MMP-8 (Z-score = 15.8) (supplemental Fig. 2 and supplemental Refs.1.and2.), MMP-8 and snapalysin (15.2), adamalysin II and ulilysin (14.4), and adamalysin II and leishmanolysin (14.3). The most distant pairs are astacin and adamalysin II (Z-score = 2.9), MMP-8 and ulilysin (6..2), and snapalysin and ulilysin (6.6).

Distinguishing Features of Each Family

Astacin is a 200-residue digestive enzyme from the crayfish A. astacus and the first member of the astacin family to be structurally analyzed (9.). Through protein degradation, growth factor activation, extracellular matrix turnover, and extracellular coat degradation (hatching), astacins participate in diverse biological processes such as digestion, development, and tissue remodeling and differentiation (e.g. promoting cartilage and bone formation and collagen biosynthesis) (27.).The three-dimensional structure of astacin shows a packman-like spherical shape and two subdomains of approximately equal size (supplemental Fig. 1A) (9.). Whereas this proteinase contains only a propeptide and a catalytic MP domain, most other astacins display additional C-terminal MATH, MAM, CUB-like, Ser/Thr-rich, I (inserted)-, epidermal growth factor-like, Tox1, and transmembrane modules (for details, see Refs.

8.,27., and28.). The astacin family has the longest region of the CSD lacking regular secondary structure elements, just a small helix and a short β-ribbon, although this is the sequentially most conserved connecting segment among metzincins (see Table 1 in Ref.7.). The protein scaffold is cross-linked by four conserved cysteine residues forming two disulfide bridges (supplemental Fig. 1). The first two residues of the mature enzyme are buried in an internal cavity in the CSD, and the N-terminal α-amino group establishes an interaction with a conserved glutamate next to the third zinc-binding histidine. The unbound coordination of the catalytic zinc is trigonal-bipyramidal due to the presence of the fourth invariant, although somewhat more distal, tyrosine zinc ligand downstream of the Met-turn (supplemental Fig. 1A). In addition to astacin, the structures of human tolloid-like protease-1 and bone morphogenetic protein-1 have recently been reported (29.).

 

Serralysins are ∼50-kDa bacterial virulence factors secreted as autoactivatable zymogens by pathogenic γ-class proteobacteria (30.). These organisms are responsible for human diseases such as meningitis, endocarditis, pyelonephritis, plague, dermatitis, soft tissue infections, septicemia, melioidosis, pneumonia, and other respiratory and urinary tract infections. They play a major role in hospital-acquired infections due to their capacity to produce surgical wound infections and to infect neonates. As part of the virulence potential of these bacteria, serralysins are directed against coagulation factors and defense-oriented proteins, protease inhibitors, lysozyme, and transferrin and may cause an anaphylactic response.

The first serralysin to be biochemically and structurally characterized was P. aeruginosa aeruginolysin (

10.). Its mature 220-residue catalytic domain lacks disulfide connections and is flanked on its C-terminal end by a calcium-stabilized β-roll domain. As in astacin, its two subdomains are of similar size. The polypeptide chain starts with an α-helix in the CSD (characteristic for the family) that is anchored to the molecular body by a conserved salt bridge with the C-terminal helix αC (supplemental Fig. 1B). The NSD features a flap made up by an elongated LβIαA and runs across the convex surface of the β-sheet. This flap varies greatly among serralysins (20 reported structures) (supplemental Table 1), distinctly affecting substrate binding. The CSD of aeruginolysin presents an extra α-helix within the segment linking the Met-turn with the wall-forming stretch and a second flap shaped by residues of the connecting segment. These elements also modulate substrate binding. Comparison of aeruginolysin, which was first solved in complex with a bound tetrapeptide (10.), with the closely related structure of unbound Serratia marcescens serralysin reveals that the unliganded zinc coordination is similar to astacin (trigonal-bipyramidal). It also includes a tyrosine that undergoes a hinge motion upon substrate binding (31.).

 

S. caespitosus snapalysin is a secreted neutral protease that comprises a 132-residue catalytic domain preceded by an alanine-rich 100-amino acid N-terminal extension including a signal peptide and a prodomain. Similar sequences have been reported for other Streptomyces species, and they have been termed SnpA (Prt and snapalysin), MprA, and SnpA. They show milk-hydrolyzing activity.

Snapalysin is the smallest metzincin and the only family member that has been structurally characterized. Its structure recalls a flattened ellipsoid and bifurcates into two asymmetric subdomains (

12.). It displays all the characteristic metzincin features, connected by short loops. Distinguishing elements are a small LβIIβIII protruding from the upper sheet within the NSD, a small bulge on top of the primed side of the active-site crevice, a short helix in LβVαB, and a calcium-binding site (supplemental Fig. 1A). In addition, an aspartate is found at the position of the third zinc-binding histidine, and two positions ahead in the sequence, a conserved tyrosine approaches but not binds the metal.

 

MMPs are secreted or membrane-bound proteinases discovered 47 years ago and participate in tail resorption during tadpole-to-frog metamorphosis. They are found mainly in higher mammals, although related sequences have been found in fish, amphibians, insects, plants, prokaryotes, and viruses. Through turnover of extracellular matrix proteins, MMPs are involved in tissue resorption, remodeling, and repair, as observed during embryogenesis and development, branching and organ morphogenesis, and angiogenesis. However, their potent proteolytic potential or its absence may also lead to pathologies such as inflammation, ulcers, rheumatoid arthritis and osteoarthritis, periodontitis, heart failure and cardiovascular disease, fibrosis, emphysema, and cancer and metastasis (32.). More recently, MMPs have been observed to be engaged in (in)activation events following limited proteolysis, as observed in apoptosis and intestinal defense protein activation but also in pathologies including stroke, human immunodeficiency virus-associated dementia, atherosclerosis, multiple sclerosis, bacterial meningitis, and Alzheimer disease. MMPs include extracellular proteins such as other (pro)proteinases, inhibitors, clotting factors, antimicrobial peptides, and chemotactic and adhesion molecules. In common with ADAMs (see below), MMPs are also involved in ectodomain shedding of growth factors, growth factor-binding proteins, hormones and hormone receptors, and cytokines and cytokine receptors from the cell surface (33.).

Like other metzincin families, MMPs are mosaic proteins constituted by a series of inserts and domains. These may include an ∼20-residue secretory signal peptide, an ∼80-residue propeptide, a 160–170-residue zinc- and calcium-dependent catalytic proteinase domain, a linker region, and a 4-fold propeller hemopexin-like C-terminal domain. Further insertions may include fibronectin type II-related domains; a collagen type V-like and vitronectin-like insertion domain; a cysteine-rich, a proline-rich, and an interleukin-1 receptor-like domain; an immunoglobulin-like domain; a glycosylphosphatidylinositol linkage signal; a membrane anchor; and a cytoplasmic tail. Naming of MMPs started historically with fibroblast collagenase as MMP-1 and has currently reached MMP-28, with 23 different forms described in humans (34.). Those MMPs encompassing a membrane anchor gave rise to the membrane-type MMP subfamily (16.,32.). Zymogen activation proceeds in MMPs according to a cysteine switch or Velcro mechanism. This removes the prodomain and switches from an inactive state, where the S^γ atom of a cysteine residue within a conserved motif, PRCGVPD, substitutes the catalytic solvent molecule in the zinc coordination sphere, to the fully accessible active enzyme (35.,36.). MMPs are the structurally most thoroughly studied metzincin family, with >120 structures reported (supplemental Table 1) (37.). The mature catalytic domain of human neutrophil collagenase (MMP-8) (13.,37.) has a shallow active-site cavity, which separates a larger NSD (∼120 residues) from a smaller CSD (∼40 amino acids), and generally a deep hydrophobic S₁′ pocket. No disulfide bonds are present in the structure. The N-terminal α-amino group is anchored to the first of two conserved aspartates imbedded in helix αC. The NSD displays an S-shaped double loop connecting strands βIII and βIV, which embraces a structural zinc cation and a tightly bound calcium ion. The downstream residues of this segment form a prominent bulge that protrudes into the active-site groove. LβIVβV and LβIIβIII contribute to a second calcium-binding site on top of the NSD β-sheet (supplemental Fig. 1, A and B). In the CSD, the MMP-8 chain displays the shortest and most conserved connecting segment within metzincins.

 

ADAMs/adamalysins/reprolysins split into three subgroups,

 the snake venom MPs, 

the mammalian ADAMs, and

 the likewise mammalian ADAMTSs (38.,39.,40.,41.). 

 The former are responsible for post-envenomation hemorrhage through digestion of extracellular matrix components surrounding capillaries, resulting in tissue necrosis. 

In turn, ADAMs were originally described to play a role in fertilization and sperm function in mammalian reproductive tracts. They are involved in myogenesis, development, neurogenesis, differentiation of osteoblastic cells, cell migration modulation, and muscle fusion. They are also engaged in human disorders like asthma, cardiac hypertrophy, obesity-associated adipogenesis and cachexia, rheumatoid arthritis, endotoxic shock, inflammation, and Alzheimer disease. They also have a major role in protein ectodomain shedding as described previously for MMPs.

 Finally, some family members lacking the transmembrane domain and harboring multiple copies of a thrombospondin 1-like repeat and a CUB domain gave rise to a distinct subfamily of soluble extracellular proteases, the ADAMTSs (42.). These enzymes disable cell adhesion by binding to integrins. They are also involved in gonad formation, embryonic development and angiogenesis, and procollagen activation, as well as in inflammatory processes, cartilage (aggrecan) degradation in arthritic diseases, bleeding disorders, and glioma tumor invasion.

All ADAMs/adamalysins/reprolysins are extracellular multidomain proteins containing a catalytic zinc- and calcium-dependent MP domain. In addition, they can display a prodomain and C-terminal disintegrin-like, cysteine-rich, C-type lectin, epidermal growth factor-like, thrombospondin 1-like, and/or transmembrane domains, as well as a cytoplasmic domain. Latency is maintained by the prodomain, and activation is believed to occur as in MMPs, i.e. by cleavage of the prodomain according to a cysteine switch-like mechanism (7.,36.,43.). 

The first catalytic domain structure to be analyzed was that of adamalysin II from C. adamanteus snake venom (11.). This is a compact 203-residue molecule of oblate ellipsoidal shape, notched at the periphery to render a relatively flat substrate-fixing cleft. This cleft separates a large ∼150-residue NSD from a small ∼50-residue CSD (supplemental Fig. 1A). Both the N and C termini are surface-located; the former is linked by a salt bridge to the C-terminal helix αC (7.). Adamalysin II deviates most from the metzincin consensus sequence within the conserved regular secondary structure elements, especially at strand βI and helices αA and αC (Fig. 1B). Inserted into the common scaffold, two additional helices are found within the NSD. In the CSD, two disulfide bonds cross-link the irregular connecting segment and attach helix αC to the NSD, respectively. A calcium ion is located on the surface, opposite the active site and close to the C terminus (supplemental Fig. 1A). The S₁′ pocket, characterized by a pronounced bulge segment LβIVβV, is hydrophobic and deep, reminiscent of some MMPs.

 In addition to adamalysin II, a number of snake venom MPs (ADAM-17, ADAM-33, and ADAMTS-1, -4, and -5) have been structurally analyzed to date (supplemental Table 1).

 

Leishmanolysins are cell-surface proteins present in most trypanosomatid, plasmodiid, and sarcocystid protozoa. They constitute the major component of the promastigote surface and are enzymatically active against polypeptide substrates. They cleave CD4 molecules at the surface of human T cells and protect promastigotes from lysis by complement proteins, suggesting a possible role as a virulence factor. Related sequences have been found in mammals (here called invadolysin), fruit flies, thale cress, nematodes, and bacteria (7.,44.). 

The only structurally analyzed family member is L. major leishmanolysin. It is synthesized as a 602-residue inactive precursor in the endoplasmic reticulum with a signal and a 100-residue propeptide, which includes a highly conserved cysteine residue potentially acting as a cysteine switch (see above). Activation liberates a mature MP of ∼280 residues, followed by an ∼200-residue C-terminal domain. A 63-residue insertion domain is observed between the glycine and the third zinc-binding histidine of the long consensus motif (supplemental Fig. 1A). The MP domain is the most asymmetric among metzincins, with a 175-residue NSD and just an ∼45-amino acid CSD. Its N terminus is located on the back left surface. The NSD is characterized by a β-sheet, which lacks strand βII (supplemental Fig. 1, A and B), and by the presence of two unique ∼40-amino acid inserted flaps, which account for most of the differences in size from the other proteins of the clan. The NSD is cross-linked by two disulfide bonds. Preceding strand βIV, a slightly prominent bulge segment lies on top of the shallow, medium-sized S₁′ pocket, which is delimited by the wall-forming segment and the beginning of the active-site helix αB. At the end of the CSD, helix αC is followed by a segment in an extended conformation, which runs from left to right across the back surface (14.).

 

The most recent family to be structurally characterized are the pappalysins. They were named after human PAPP-A, a heavily glycosylated 170-kDa multidomain protein specifically cleaving insulin-like growth factor-binding proteins (45.). Proulilysin is a 38-kDa archaeal protein from M. acetivorans that shares sequence similarity with PAPP-A, but it encompasses only the prodomain and the catalytic domain (15.). The proprotein may undergo cysteine switch-mediated activation, as suggested by the presence of a conserved cysteine in the prodomain. Activation occurs autolytically in the presence of calcium. 

 

With 262 residues, mature ulilysin is the largest MP of all metzincin catalytic domains. As distinguishing features, it presents in the NSD a loop dividing strand βII into two substrands (βII and βII′) and a β-ribbon inserted within LβIIIβIV that protrudes from the molecular surface and frames the active site on its primed side. The segment connecting helix αA with strand βII is the largest among metzincins and covers almost all of the back of the molecule from the NSD to the CSD in a cape-like fashion and includes two unique α-helices. The glycine of the zinc-binding motif is replaced in ulilysin and a small subset of pappalysins by an asparagine under slight variation of the main-chain angles, which do not correspond here to a high-energy conformation but to a left-handed α-helix. Overall, the chain trace flanking this residue is indistinguishable from other metzincins (supplemental Fig. 1) (7.). The CSD shows two disulfide bonds and a unique two-calcium site. This site is a molecular switch for activity, as the proteinase can be reversibly inhibited through calcium chelators ( 15.,20.,21.). Finally, in the absence of an unbound structure, ulilysin may possess a fifth zinc-binding tyrosine ligand provided by the Met-turn that is swung out upon substrate binding.

Conclusions

The metzincins constitute a clan of ubiquitous MPs present in all kingdoms of life, which are pivotal for physiology and pathology. So far, seven families have been structurally analyzed. The clan represents a case of divergent evolution from an urmetzincin, which may not look very different from the smallest member, snapalysin. Into such a minimal scaffold, evolution has introduced unique structural elements conserved among each family. Based on the presence of an extended zinc-binding consensus sequence pattern, several more families have been suggested to enlarge the clan, although future structural analysis will be required to confirm their ascription. Structural information on each of the seven families may help to elucidate common catalytic and processing mechanisms and to assign the function of proteins encoded by newly discovered gene sequences.

Acknowledgments

I thank Robin Rycroft for help

COVID-19 ja ADAMTS-19

 https://www.practiceupdate.com/content/adamts13-regulation-of-vwf-multimer-distribution-in-severe-covid-19/119629

ADAMTS13 ja vWf. Hematologiset taudit iTTP ja hTTP.Tromboottiset trombosytopeniset purpuramuodot.

 Löydän tänään tulleesta tanskalaisesta lääkärilehdestä hematologia käsittelevien artikkeleiden joukosta  myös ADAMTS13 geeniä käsittelevää. 

Hansen,A C Nilsson, Henrik Fredriksen. Trombotisk trombocytopeniskt purpura TTP tai TMA.

TTP on disseminoitunutta mikrotrombimuodostusta luonteeltaan. TTP on akuutti, henkeä uhkaava tromboottinen mikroangiopatia. Tilaa aiheuttaa ADAMTS13- entsyymin puute.

Tieteen edistyminen  on lähinnä tilanteen arvioinnin ja täsmällisen  biokemiallisen diagnostiikan ja  hoidon  optimoitumista.

Seuranta vaatii ADAMTS13-entsyymiaktiiviteetin monitorointia. Myös psykiatriset ja kognitiiviset  seuraamuksetkin on otettava huomioon.

Taudilla on kaksi muotoa iTTP ja hTTP

iTTP,  hankittu  autoimmuunitauti, jossa immunoglobuliini G-autovasta-aineet kohdistuvat ADAMTS13-proteaasientsyymiin, joilloin siitä tulee puutetta tavalla tai toisella: joko entsyymin aktiivisuus estyy tai sen vaihtuminen kiihtyy.

hTTP, hereditäärinen, perinnöllinen  tautimuoto johtunee resessiivisesti periytyvistäå mutatoituneista ADAMTS13 geeneistä ADAMTS13-entsyymin aktiivisuus on tällöin alentunutta.  Tämä  hTTP on harvinaista. Sitä on 5%:ssa kaikista TTP-tapauksista. Naisilla esiintyy useammin TTP.

Kertausta  ADAMTS13 proteaasin  fysiologisesta tehtävästä:

ADAMTS13 pilkkoo  vW-faktorin molekyylejä, kun niitä  alkaa ultrapitkinä  vapautua endoteelisoluista ja saa aikaan monomeerimuotoja.

ADAMTS13- entsyymin puutteessa jää vW-faktorista pitkiä  muotoja , jotka takerruttavat  verihiutaleita toisiinsa  aiheuttaen  mikrotrombeja. Tämä  voi tapahtua myös juoksevassa  veressä jossa ei ole  paikallisvauriota. Siis ilmiö voi olla disseminoitua.Koska nyt  aggrekoituu  trombosyyttejä, niistä tulee  puutetta vähitellen: konsumptiosytopenia , lisäksi vielä punasolujen mekaanista vaurioitumista Kliinisesti tilanne ilmenee hemolyyttisenä anemiana ja trombosytopeniana ja fluktuoivina   elinvaikutuksina, joista heijastuu mikrotrombien muodostuminen ja  liukeneminen. 

Kolmasosalla potilaista esiintyy myös neurologisia oireita.päänsärkyä,  TIA-oiretta,jopa halvausta, koko aivokuoren antamaa affektiivista oireilua tai tajuttomuutta. Kolmasosalla esiintyy kuumetta, pahoinvointia ja mahakipuja.  Munuaisfunktiossa voi olla vaikutusta n 40%:lla, useimmiten  vain lievästi kohonneita kreatiniinipitoisuuksia. Kuitenkin dialyysitarvetta on  1-2%:lla. 

60%:lla iTTP -potilaista ei esiinny  toisen tekijän  liittymistä tilaan, Useimmiten assosioituvia ovat jokin toinen autoimmuniteetti, infektio   tai raskaudentila. 

Verenkuvan piirteitä TTP-potilailla:

Aina trombosytopenia ja merkkejä mikroangiopaattisesta hemolyyttisestä anemiasta, jossa on matala hemoglobiinipitoisuus,matala haptoglobiinitaso, korkea retikulosyyttipitoisuus, korkea  vapaa hemoglobiini-, bilirubiini-ja laktaattidehydrogenaasipitoisuus sekä lisääntynyt määrä fragmentoituneita punasoluja(skistosyyttejä).

Koagulaatiotekijät ovat normaaleja ja suora antiglobuliinitesti on negatiivinen. ADAMTS13- entsyymipitoisuudet ovat alle 10% normaalista. (ref.40-130%)  iTTP-tapauksissa todetaan myös ADAMTS13- autovasta-aineita. 

Elinvaikutus voi heijastua esim troponiini ja kreatiniinipitoisuuksien kohoamisissa.

CT/MR on tyypillisesti normaali komatoosissakin  tilassa.

Hoito:Plasmanvaihto, jolla vähennetään  ADAMTS13- autovasta-aineita. ADAMTS13- entsyymin anto.  Kortikosteroideja, Rituximab, Caplacizumab.

ugeskriftet.dk 18.oktober 2021

Ugeskr Laeger2021;183;V03210230

Jänteen kehityksestä. Tenosyyttisolut. Metalloproteinaasien MMP ja ADAMTS metalloproteaasien osuudesta.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4689213/

 

 

 

Covid-19 ja  nivelet, jänteet?

Col6A1

Interactome  Sars-2 , Human trofoblast 

file:///C:/Users/Laatikko/AppData/Local/Temp/preprints202005.0333.v1.pdf