Biology:Coagulin

From HandWiki
Revision as of 03:53, 12 February 2024 by StanislovAI (talk | contribs) (correction)
(diff) ← Older revision | Latest revision (diff) | Newer revision → (diff)
Coagulin
Identifiers
SymbolCoagulin
PfamPF02035
InterProIPR000275
SCOP2d1aoca_ / SCOPe / SUPFAM
Limulus polyphemus horseshoe crabs use coagulin to form gel clots

Coagulin is a gel-forming protein of hemolymph that hinders the spread of bacterial and fungal invaders by immobilizing them. It is produced in the coagulogen form before being cleaved into the active form through a serine proteinase cascade.[1][2][3] It has been most extensively studied in horseshoe crabs. It has also been produced by other organisms, such as Bacillus coagulans I4 in a plasmid location.[4] In human medicine, coagulation of coagulin is the basis of detection of bacterial endotoxin through the Limulus amebocyte lysate test for parenteral medications.

Structure

Coagulogen contains a single 175-residue polypeptide chain that is cleaved after Arg-18 and Arg-46 by a Limulus clotting enzyme contained in the granular hemocyte cells of the hemolymph. A pathway is initiated in which ultimately the limulus clotting enzyme cleaves coagulogen to coagulin. Cleavage releases two chains of coagulin, chains A and B, covalently linked by two disulfide bonds, together with the peptide C.[1][2][5] The A-B fold wraps around the helical peptide C, forming a compact structure.[6] The approximate mass of coagulin is 3-4 kDA by SDS-PAGE.[4] Gel formation results from interlinking of coagulin molecules.[1] Before interlinking the coagulin monomers, peptide C is cleaved from coagulogen. Removal of peptide C exposes an extended hydrophobic cove of the newly cleaved molecule, allowing interaction with a second molecule’s hydrophobic edge.[6][7] The full-length structure of a coagulogen is known (PDB: 1AOC​); it shares the same cystine-knot cytokine superfamily (fold) as neurotrophins, with several cystines conserved.

Coagulation

Hemolymph coagulation is a part of the invertebrate immune response. Factors within the hemolymph are activated and initiate a pathway where insoluble clots are formed in order to prevent leakage of bodily fluids and immobilized microbes from infecting the organism.[8] This is crucial as invertebrate organisms do not have adaptive immune systems comparable to the one in the mammalian immune system.[7] In crustaceans, hemolymph coagulation depends on the transglutaminase-mediated cross-linking of specific plasma-clotting proteins, but without the proteolytic cascade.[9]

In horseshoe crabs, the proteolytic coagulation cascade is triggered by lipopolysaccharides and beta-1,3-glucans. There are two types of hemocytes within the horseshoe crab hemolymph: granular and nongranular. The granular hemocytes are activated by bacterial endotoxins lipopolysaccharides (LPS) that are found on the surface of Gram-negative bacteria. “...LPS comprises approximately 70% of the outer membranes of gram-negative bacteria.”[10] They are also activated by beta-1,3-glucans that are found on the cell walls of yeast and some fungi.[5][7]

In the LPS-activated pathway, LPS activates zymogen factor C. It is autocatalytically converted into the activated form factor C. The active factor C converts inactive factor B into active factor B. Active factor B converts the proclotting enzyme into the clotting enzyme.[5] The clotting enzyme cleaves coagulogen into coagulin, resulting in noncovalent coagulin homopolymers through head-to-tail interaction.

In the beta-1,3-glucan activated pathway, there are slight differences. Beta-1,3-glucan activates zymogen factor G. It is autocatalytically converted into the activated form factor G. From here, the pathway converges into the LPS-activated pathway. The active factor G converts the proclotting enzyme into the clotting enzyme to cleave coagulogen into coagulin.[5] In both pathways, gel formation occurs when the final enzyme transglutaminase cross-links coagulin.[7]

However, horseshoe crab transglutaminase does not cross-link coagulins intermolecularly. Recently, coagulins were discovered to be cross-linked on hemocyte cell surface proteins called proxins. This indicates that a cross-linking reaction at the final stage of hemolymph coagulation is an important innate immune system of horseshoe crabs.[9]

In comparison, mammalian blood coagulation differs from hemolymph coagulation. Mammalian blood coagulation is largely dependent on platelets and fibrin, whereas hemolymph does not contain platelets or fibrin but hemocytes.[11] Mammalian blood coagulation is based on the proteolytically induced polymerization of fibrinogens. There are two pathways (Tissue factor and Contact) that result in thrombin converting fibrinogen to fibrin. Fibrin monomers noncovalently interact with each other and polymerize to form the blood clot.[12] Fibrin and coagulin are analogous to each other. Similarities between mammalian blood coagulation and hemolymph coagulation include gel formation, TGase, and serve as a part of wound healing.[7] However, the clot formed in hemolymph coagulation is softer than the mammalian fibrin clot.[5][7]

Uses

Limulus amebocyte lysate test

Limulus amebocyte lysate is found only in horseshoe crabs, specifically the Limulus polyphemus species. In the presence of bacterial endotoxins (LPS) and beta-1,3-glucans, it initiates the coagulation pathways . It is employed as an FDA-approved assay method to test sterility of medical instruments and injectable drugs, such as in the pharmaceutical industry.[13]

Since the 1970s, the Limulus amebocyte lysate test has been used to test for endotoxins in human blood samples. The original method (Limulus gelation test) involved qualitatively looking for coagulin gel formation. After a one hour incubation, if the sample was coagulated, it formed a solid clot that was positive for endotoxins. If the sample was not coagulated, it would be liquid and was negative for endotoxins.[10] However, the technique was limited by its sensitivity.[14]

Today, the Limulus test is one hundred times more sensitive and uses a chromogenic method of detection.[5] When coagulogen is cleaved by the clotting enzyme, coagulin is produced. However the clotting enzyme also produces a chromogenic end product known as pNA. pNA (Boc-Leu-Gly-Arg-p-nitroanilide) is the chromogenic product that emits a yellow color.[15][16] The concentration of endotoxins in a sample can be calculated by measuring the absorbance of released pNA at 405 nm.[5][16]

Evolution

Coagulin is found in the four species of horseshoe crabs: Limulus polyphemus, Tachypleus tridentatus, Tachypleus gigas, and Carcinoscorpius rotundicauda. They are deemed “living fossils” as they have been around for 445-500 million years with little significant change compared to their ancestors.[7][13] The coagulin precursor, coagulogen, has a mutation rate of 1.2 x 10-9 per amino acid per year as compared to its mammalian analog, fibrinogen, with a mutation rate of 8.3 x 10-9.[7] It is contained in hemocytes, a type of phagocyte. There are different types of phagocytes and are found in all invertebrate groups (as either hemocytes, amoebocytes, or coelomocytes). Comparing vertebrae and Limulus polyphemus coagulation systems, none of the cascade proteins (including coagulogen) share a common protein domain with two exceptions, Hemolectin and TGase. While the two systems are functionally similar, the coagulation proteins “have different evolutionary histories.”[17]

See also

References

  1. ↑ 1.0 1.1 1.2 "The complete amino acid sequence of coagulogen isolated from Southeast Asian horseshoe crab, Carcinoscorpius rotundicauda". Journal of Biochemistry 98 (2): 305–318. August 1985. doi:10.1093/oxfordjournals.jbchem.a135283. PMID 3905780. 
  2. ↑ 2.0 2.1 "The amino acid sequence of coagulogen isolated from southeast Asian horseshoe crab, Tachypleus gigas". Journal of Biochemistry 95 (6): 1793–1801. June 1984. doi:10.1093/oxfordjournals.jbchem.a134792. PMID 6469947. 
  3. ↑ "Coagulation, an ancestral serine protease cascade, exerts a novel function in early immune defense". Blood 118 (9): 2589–2598. September 2011. doi:10.1182/blood-2011-02-337568. PMID 21613262. 
  4. ↑ 4.0 4.1 "Coagulin, a bacteriocin-like inhibitory substance produced by Bacillus coagulans I4". Journal of Applied Microbiology 85 (1): 42–50. July 1998. doi:10.1046/j.1365-2672.1998.00466.x. PMID 9721655. 
  5. ↑ 5.0 5.1 5.2 5.3 5.4 5.5 5.6 "Biochemical principle of Limulus test for detecting bacterial endotoxins". Proceedings of the Japan Academy. Series B, Physical and Biological Sciences 83 (4): 110–119. May 2007. doi:10.2183/pjab.83.110. PMID 24019589. Bibcode: 2007PJAB...83..110I. 
  6. ↑ 6.0 6.1 "Crystal structure of a coagulogen, the clotting protein from horseshoe crab: a structural homologue of nerve growth factor". The EMBO Journal 15 (24): 6789–6797. December 1996. doi:10.1002/j.1460-2075.1996.tb01070.x. PMID 9003754. 
  7. ↑ 7.0 7.1 7.2 7.3 7.4 7.5 7.6 7.7 "Evolution and phylogeny of defense molecules associated with innate immunity in horseshoe crab". Frontiers in Bioscience 3 (4): D973–D984. September 1998. doi:10.2741/A337. PMID 9727083. 
  8. ↑ "Hemolymph coagulation". QuickGO. European Bioinformatics Institute. https://www.ebi.ac.uk/QuickGO/GTerm?id=GO:0042381. 
  9. ↑ 9.0 9.1 "Structure and function of coagulogen, a clottable protein in horseshoe crabs". Cellular and Molecular Life Sciences 61 (11): 1257–1265. June 2004. doi:10.1007/s00018-004-3396-5. PMID 15170505. 
  10. ↑ 10.0 10.1 "Horseshoe Crab Aquaculture as a Sustainable Endotoxin Testing Source". Frontiers in Marine Science 7. 2020-04-01. doi:10.3389/fmars.2020.00153. ISSN 2296-7745. 
  11. ↑ "Explore the Differences between Blood and Haemolymph" (in en). https://byjus.com/biology/difference-between-blood-and-haemolymph/. 
  12. ↑ "How it all starts: Initiation of the clotting cascade". Critical Reviews in Biochemistry and Molecular Biology 50 (4): 326–336. 2015-07-04. doi:10.3109/10409238.2015.1050550. PMID 26018600. 
  13. ↑ 13.0 13.1 "Facts About Horseshoe Crabs and FAQ" (in en). https://myfwc.com/research/saltwater/crustaceans/horseshoe-crabs/facts/. 
  14. ↑ "Limulus amebocyte lysate (LAL) detection of endotoxin in human blood" (in en). Journal of Endotoxin Research 1 (4): 253–263. December 1994. doi:10.1177/096805199400100407. ISSN 0968-0519. 
  15. ↑ "Bacterial Endotoxin test". Wako LAL System. https://labchem-wako.fujifilm.com/asia/lal/lal_knowledge/about_lal_measurement.html. 
  16. ↑ 16.0 16.1 "Endotoxin and pyrogen testing". Pharmaceutical Microbiology. Elsevier. 2016. pp. 131–145. doi:10.1016/b978-0-08-100022-9.00011-6. ISBN 9780081000229. 
  17. ↑ "Domain Evolution of Vertebrate Blood Coagulation Cascade Proteins". Journal of Molecular Evolution 90 (6): 418–428. December 2022. doi:10.1007/s00239-022-10071-3. PMID 36181519. Bibcode: 2022JMolE..90..418C. 
This article incorporates text from the public domain Pfam and InterPro: IPR000275



Retrieved from "https://handwiki.org/wiki/index.php?title=Biology:Coagulin&oldid=3586541"