Summary: A thermostable extracellular metalloendopeptidase containing four calcium ions. (Enzyme Nomenclature, 1992)

Top Publications

  1. Kin T, Zhai X, Murdoch T, Salam A, Shapiro A, Lakey J. Enhancing the success of human islet isolation through optimization and characterization of pancreas dissociation enzyme. Am J Transplant. 2007;7:1233-41 pubmed
    ..Our results from 251 islet isolations showed that optimization of thermolysin dosage based on Caseinase unit/g pancreas contributed considerably to islet isolation outcome but that ..
  2. Park C, Marqusee S. Probing the high energy states in proteins by proteolysis. J Mol Biol. 2004;343:1467-76 pubmed
    ..Mass spectrometry and N-terminal sequencing showed that thermolysin cleaves the peptide bond between Thr92 and Ala93 in an extended loop region of the protein...
  3. Rosengren K, Blond A, Afonso C, Tabet J, Rebuffat S, Craik D. Structure of thermolysin cleaved microcin J25: extreme stability of a two-chain antimicrobial peptide devoid of covalent links. Biochemistry. 2004;43:4696-702 pubmed
    ..of a two-chain peptide formed by the treatment of the potent antimicrobial peptide microcin J25 (MccJ25) with thermolysin has been characterized by NMR spectroscopy and mass spectrometry...
  4. Gragnani A, Sobral C, Ferreira L. Thermolysin in human cultured keratinocyte isolation. Braz J Biol. 2007;67:105-9 pubmed
    ..To compare the action of trypsin and thermolysin in the keratinocyte isolation using newborn foreskin...
  5. Trusek Holownia A. Synthesis of ZAlaPheOMe, the precursor of bitter dipeptide in the two-phase ethyl acetate-water system catalysed by thermolysin. J Biotechnol. 2003;102:153-63 pubmed
    ..The selected system and process conditions (60 degrees C, pH 7.0) ensured high activity and stability of thermolysin, a catalyst of this reaction...
  6. Miyoshi S, Nakazawa H, Kawata K, Tomochika K, Tobe K, Shinoda S. Characterization of the hemorrhagic reaction caused by Vibrio vulnificus metalloprotease, a member of the thermolysin family. Infect Immun. 1998;66:4851-5 pubmed
    ..Therefore, specific degradation of type IV collagen may cause destruction of the basement membrane, breakdown of capillary vessels, and leakage of blood components including erythrocytes. ..
  7. O Gorman D, Kin T, Imes S, Pawlick R, Senior P, Shapiro A. Comparison of human islet isolation outcomes using a new mammalian tissue-free enzyme versus collagenase NB-1. Transplantation. 2010;90:255-9 pubmed publisher
    ..We report our experience using the MTF enzyme in clinical islet isolations compared with Serva NB-1 with modified enzyme delivery method...
  8. Ubl J, Grishina Z, Sukhomlin T, Welte T, Sedehizade F, Reiser G. Human bronchial epithelial cells express PAR-2 with different sensitivity to thermolysin. Am J Physiol Lung Cell Mol Physiol. 2002;282:L1339-48 pubmed
    ..Cathepsin G altered neither the resting Ca(2+) level nor PAR-2-elicited Ca(2+) response. Thermolysin, a prototypic bacterial metalloprotease, induced a dose-dependent Ca(2+) response in HBE, but not A549, cells...
  9. Mungall B, Pollitt C. Thermolysin activates equine lamellar hoof matrix metalloproteinases. J Comp Pathol. 2002;126:9-16 pubmed
    ..Explants treated with the bacterial protease thermolysin separated dose-dependently; this was accompanied by activation of both MMP-2 and -9...

More Information


  1. Papadopoulos T, Kelly J, Bauer K. Mutational analysis of the thyrotropin-releasing hormone-degrading ectoenzyme. similarities and differences with other members of the M1 family of aminopeptidases and thermolysin. Biochemistry. 2001;40:9347-55 pubmed
    ..of TRH-DE while E464 presumably represents the third zinc-coordinating residue and may be equivalent to E166 in thermolysin. In contrast, amino acid residues R488 and Y554 seem not to be involved in the catalytic mechanism of TRH-DE.
  2. Selkti M, Tomas A, Gaucher J, Prangé T, Fournie Zaluski M, Chen H, et al. Interactions of a new alpha-aminophosphinic derivative inside the active site of TLN (thermolysin): a model for zinc-metalloendopeptidase inhibition. Acta Crystallogr D Biol Crystallogr. 2003;59:1200-5 pubmed
    ..This could be taken into account in the design of selective inhibitors. ..
  3. Khan M, Fuskevåg O, Sylte I. Discovery of potent thermolysin inhibitors using structure based virtual screening and binding assays. J Med Chem. 2009;52:48-61 pubmed publisher
    In the present work, 22 compounds of the U.S. NCI compound library (size 273K) were identified as putative thermolysin binders by structure based virtual screening with the ICM software (ICM-VLS)...
  4. Adekoya O, Sylte I. The thermolysin family (M4) of enzymes: therapeutic and biotechnological potential. Chem Biol Drug Des. 2009;73:7-16 pubmed publisher
    ..Some enzymes of the family are able to function at the extremes of temperatures, and some function in organic solvents. Thereby enzymes of the thermolysin family have an innovative potential for biotechnological applications.
  5. Juers D, Kim J, Matthews B, Sieburth S. Structural analysis of silanediols as transition-state-analogue inhibitors of the benchmark metalloprotease thermolysin. Biochemistry. 2005;44:16524-8 pubmed
    ..the design of active-site-directed protease inhibitors, including aspartic (HIV protease) and metallo (ACE and thermolysin) proteases...
  6. Hernández Ledesma B, Ramos M, Recio I, Amigo L. Effect of beta-lactoglobulin hydrolysis with thermolysin under denaturing temperatures on the release of bioactive peptides. J Chromatogr A. 2006;1116:31-7 pubmed
    In this study, bovine beta-lactoglobulin A (beta-Lg A) was hydrolysed with thermolysin under non-denaturing and heat-denaturing conditions. The peptides released during hydrolysis were identified by HPLC-MS/MS...
  7. Owen J, Maddison B, Whitelam G, Gough K. Use of thermolysin in the diagnosis of prion diseases. Mol Biotechnol. 2007;35:161-70 pubmed
    ..Here, we have applied the protease thermolysin to the diagnosis of animal prion diseases...
  8. Takita T, Aono T, Sakurama H, Itoh T, Wada T, Minoda M, et al. Effects of introducing negative charges into the molecular surface of thermolysin by site-directed mutagenesis on its activity and stability. Biochim Biophys Acta. 2008;1784:481-8 pubmed publisher
    b>Thermolysin is remarkably activated and stabilized by neutral salts, and surface charges are suggested important in its activity and stability...
  9. Demidyuk I, Kalashnikov A, Gromova T, Gasanov E, Safina D, Zabolotskaya M, et al. Cloning, sequencing, expression, and characterization of protealysin, a novel neutral proteinase from Serratia proteamaculans representing a new group of thermolysin-like proteases with short N-terminal region of precursor. Protein Expr Purif. 2006;47:551-61 pubmed
    ..The gene encoded a precursor of 341 amino acids (AAs) with a significant homology to thermolysin-like proteinases (TLPs)...
  10. Kin T, O Gorman D, Zhai X, Pawlick R, Imes S, Senior P, et al. Nonsimultaneous administration of pancreas dissociation enzymes during islet isolation. Transplantation. 2009;87:1700-5 pubmed publisher
    ..This beneficial effect of the modified method was, however, diminished when applied to younger donor pancreata. Our study brings new insight into the role of collagenase and noncollagenolytic protease on pancreas dissociation. ..
  11. Kim J, Sieburth S. A silanediol inhibitor of the metalloprotease thermolysin: synthesis and comparison with a phosphinic acid inhibitor. J Org Chem. 2004;69:3008-14 pubmed
    A silanediol inhibitor of the metalloprotease thermolysin was prepared for comparison to a known phosphinic acid inhibitor, providing the first comparison of these second-row element based transition-state analogues...
  12. English A, Groom C, Hubbard R. Experimental and computational mapping of the binding surface of a crystalline protein. Protein Eng. 2001;14:47-59 pubmed
    ..We determined the high-resolution crystal structures of thermolysin (TLN), generated from crystals soaked in 50--70% acetone, 50--80% acetonitrile and 50 mM phenol...
  13. Murray C, Baxter C, Frenkel A. The sensitivity of the results of molecular docking to induced fit effects: application to thrombin, thermolysin and neuraminidase. J Comput Aided Mol Des. 1999;13:547-62 pubmed
    ..All-pairs docking experiments are performed for three enzymes (thrombin, thermolysin and influenza virus neuraminidase) based on six or more ligand-enzyme crystal structures for each enzyme...
  14. Demidyuk I, Gasanov E, Safina D, Kostrov S. Structural organization of precursors of thermolysin-like proteinases. Protein J. 2008;27:343-54 pubmed publisher
    The primary structures of the full-length precursors of thermolysin-like proteinases (TLPs) were systemically analyzed...
  15. Miyoshi S, Sonoda Y, Wakiyama H, Rahman M, Tomochika K, Shinoda S, et al. An exocellular thermolysin-like metalloprotease produced by Vibrio fluvialis: purification, characterization, and gene cloning. Microb Pathog. 2002;33:127-34 pubmed
    ..protease, including a molecular mass of 45kDa, sensitivity to chelating agents or competitive inhibitors for thermolysin-like metalloproteases, and the substrate specificity...
  16. Kim J, Glekas A, McN Sieburth S. Silanediol-based inhibitor of thermolysin. Bioorg Med Chem Lett. 2002;12:3625-7 pubmed
    The first silanediol inhibitor of thermolysin is reported, prepared by analogy with the Grobelny/Bartlett phosphinate inhibitor...
  17. Owen J, Rees H, Maddison B, Terry L, Thorne L, Jackman R, et al. Molecular profiling of ovine prion diseases by using thermolysin-resistant PrPSc and endogenous C2 PrP fragments. J Virol. 2007;81:10532-9 pubmed
    ..Here we describe a novel molecular strain typing assay that used thermolysin digestion of caudal medulla samples to produce PrPres signatures on Western blots that readily distinguished ..
  18. Xiao Y, Morice A, Compton S, Sadofsky L. N-linked glycosylation regulates human proteinase-activated receptor-1 cell surface expression and disarming via neutrophil proteinases and thermolysin. J Biol Chem. 2011;286:22991-3002 pubmed publisher
    ..of hPAR(1) downstream of the tethered ligand (especially Asn(75)) governs receptor disarming to trypsin, thermolysin, and the neutrophil proteinases elastase and proteinase 3 but not cathepsin G...
  19. Park C, Marqusee S. Pulse proteolysis: a simple method for quantitative determination of protein stability and ligand binding. Nat Methods. 2005;2:207-12 pubmed
    ..The simplicity of pulse proteolysis suggests that it is an excellent strategy for the high-throughput determination of protein stability in protein engineering and drug discovery applications. ..
  20. Pelmenschikov V, Blomberg M, Siegbahn P. A theoretical study of the mechanism for peptide hydrolysis by thermolysin. J Biol Inorg Chem. 2002;7:284-98 pubmed
    The catalytic mechanism for peptide hydrolysis by thermolysin has been investigated using the B3LYP hybrid density functional method...
  21. Inouye K, Kuzuya K, Tonomura B. Sodium chloride enhances markedly the thermal stability of thermolysin as well as its catalytic activity. Biochim Biophys Acta. 1998;1388:209-14 pubmed
    b>Thermolysin, a thermophilic metalloproteinase, is markedly activated in the presence of high concentrations (1-5 M) of neutral salts...
  22. Mock W, Stanford D. Arazoformyl dipeptide substrates for thermolysin. Confirmation of a reverse protonation catalytic mechanism. Biochemistry. 1996;35:7369-77 pubmed
    Cleavage by thermolysin of N-(4-methoxyphenylazoformyl)-L-leucyl-L-leucine plus some congeneric peptides provides a highly sensitive new kinetic assay for proteolytic activity...
  23. Kuzuya K, Inouye K. Effects of cobalt-substitution of the active zinc ion in thermolysin on its activity and active-site microenvironment. J Biochem. 2001;130:783-8 pubmed
    b>Thermolysin is remarkably activated in the presence of high concentrations (1-5 M) of neutral salts [Inouye, K. (1992) J. Biochem. 112, 335-340]. The activity is enhanced 13-15 times with 4 M NaCl at pH 7.0 and 25 degrees C...
  24. Balamurugan A, Loganathan G, Bellin M, Wilhelm J, Harmon J, Anazawa T, et al. A new enzyme mixture to increase the yield and transplant rate of autologous and allogeneic human islet products. Transplantation. 2012;93:693-702 pubmed publisher
    ..The ECs consisted of purified, intact or truncated class 1 (C1) and class 2 (C2) collagenases from Clostridium histolyticum (Ch), and neutral protease (NP) from Bacillus thermoproteolyticus rokko (thermolysin) or Ch (ChNP).
  25. Rakhorst H, Tra W, Posthumus van Sluijs S, de Groot E, van Osch G, van Neck J, et al. Mucosal keratinocyte isolation: a short comparative study on thermolysin and dispase. Int J Oral Maxillofac Surg. 2006;35:935-40 pubmed
    ..The purpose of this study was to compare dispase and thermolysin for keratinocyte isolation...
  26. Inouye K, Kusano M, Hashida Y, Minoda M, Yasukawa K. Engineering, expression, purification, and production of recombinant thermolysin. Biotechnol Annu Rev. 2007;13:43-64 pubmed
    b>Thermolysin [EC] is a thermostable neutral zinc metalloproteinase originally identified in the culture broth of Bacillus thermoproteolyticus Rokko...
  27. Brandhorst H, Friberg A, Andersson H, Felldin M, Foss A, Salmela K, et al. The importance of tryptic-like activity in purified enzyme blends for efficient islet isolation. Transplantation. 2009;87:370-5 pubmed publisher
    ..Previous studies have solely focused on the presence of collagenase and neutral protease/thermolysin. Despite improved characterization of these components, the lot-related variability in efficacy still persists ..
  28. Kim M, Song J, Park C. Determining protein stability in cell lysates by pulse proteolysis and Western blotting. Protein Sci. 2009;18:1051-9 pubmed publisher
    ..This method allows the investigation of conformational energetics of proteins in cell lysates without cloning, purification, or labeling. ..
  29. Cronier S, Gros N, Tattum M, Jackson G, Clarke A, Collinge J, et al. Detection and characterization of proteinase K-sensitive disease-related prion protein with thermolysin. Biochem J. 2008;416:297-305 pubmed publisher
    ..Recently, thermolysin has been identified as a complementary tool to PK, permitting isolation of PrP(Sc) in its full-length form...
  30. Liu Z, Zhang P, Zhou Y, Qin H, Shen T. Culture of human intestinal epithelial cell using the dissociating enzyme thermolysin and endothelin-3. Braz J Med Biol Res. 2010;43:451-9 pubmed
    ..b>Thermolysin was added to improve the yield of epithelial cells, while endothelin-3 was added to stimulate their growth...
  31. Xie B, Bian F, Chen X, He H, Guo J, Gao X, et al. Cold adaptation of zinc metalloproteases in the thermolysin family from deep sea and arctic sea ice bacteria revealed by catalytic and structural properties and molecular dynamics: new insights into relationship between conformational flexibility and. J Biol Chem. 2009;284:9257-69 pubmed publisher
    ..We reported two novel cold-adapted zinc metalloproteases in the thermolysin family, vibriolysin MCP-02 from a deep sea bacterium and vibriolysin E495 from an Arctic sea ice bacterium, and ..
  32. Englert L, Silber K, Steuber H, Brass S, Over B, Gerber H, et al. Fragment-based lead discovery: screening and optimizing fragments for thermolysin inhibition. ChemMedChem. 2010;5:930-40 pubmed publisher
    ..of in silico fragment-based screening for the discovery of novel lead compounds for the metalloendoproteinase thermolysin. We have chosen thermolysin to validate our screening approach as it is a well-studied enzyme and serves as a ..
  33. Young T, Skordalakes E, Marqusee S. Comparison of proteolytic susceptibility in phosphoglycerate kinases from yeast and E. coli: modulation of conformational ensembles without altering structure or stability. J Mol Biol. 2007;368:1438-47 pubmed
    ..coli PGK, while the characteristics defining the yeast homolog suggest that proteolysis occurs upon unfolding of only the C-domain, with the N-domain remaining folded and consequently resistant to cleavage. ..
  34. Berney T, Molano R, Cattan P, Pileggi A, Vizzardelli C, Oliver R, et al. Endotoxin-mediated delayed islet graft function is associated with increased intra-islet cytokine production and islet cell apoptosis. Transplantation. 2001;71:125-32 pubmed
    ..This emphasizes the fact that using endotoxin-free reagents during isolation is a key factor for successful islet transplantation. ..
  35. Ragusa S, Mouchet P, Lazennec C, Dive V, Meinnel T. Substrate recognition and selectivity of peptide deformylase. Similarities and differences with metzincins and thermolysin. J Mol Biol. 1999;289:1445-57 pubmed
    ..Based on the numerous homologies that deformylase displays with thermolysin and metzincins, a mechanism of enzyme:substrate recognition and hydrolysis could finally be proposed...
  36. Murphy L, Corder R, Mallet A, Turner A. Generation by the phosphoramidon-sensitive peptidases, endopeptidase-24.11 and thermolysin, of endothelin-1 and c-terminal fragment from big endothelin-1. Br J Pharmacol. 1994;113:137-42 pubmed
    1. Phosphoramidon, a potent inhibitor of endopeptidase-24.11 (E-24.11) and thermolysin, has been shown to reduce the hypertensive effect of exogenous big endothelin-1 (big ET-1) in rats. To examine whether E-24...
  37. Holland D, Tronrud D, Pley H, Flaherty K, Stark W, Jansonius J, et al. Structural comparison suggests that thermolysin and related neutral proteases undergo hinge-bending motion during catalysis. Biochemistry. 1992;31:11310-6 pubmed
    Crystal structures are known for three members of the bacterial neutral protease family: thermolysin from Bacillus thermoproteolyticus (TLN), the neutral protease from Bacillus cereus (NEU), and the elastase of Pseudomonas aeruginosa (..
  38. Savitz A, Meyer D. Identification of a ribosome receptor in the rough endoplasmic reticulum. Nature. 1990;346:540-4 pubmed
    ..Isolation of this domain has led to the identification, purification and characterization of the intact ribosome receptor, as well as its functional reconstitution into lipid vesicles. ..
  39. Preusser Kunze A, Mariappan M, Schmidt B, Gande S, Mutenda K, Wenzel D, et al. Molecular characterization of the human Calpha-formylglycine-generating enzyme. J Biol Chem. 2005;280:14900-10 pubmed
    ..Peptides derived from all known human sulfatases served as substrates for purified FGE indicating that FGE is sufficient to modify all sulfatases of the same species. ..
  40. Hopkinson A, Shanmuganathan V, Gray T, Yeung A, Lowe J, James D, et al. Optimization of amniotic membrane (AM) denuding for tissue engineering. Tissue Eng Part C Methods. 2008;14:371-81 pubmed publisher
    ..denuded using published ethylenediaminetetraacetic acid (EDTA)- and dispase-based methodologies and our novel thermolysin-based procedure...
  41. Kim S, Kim D, Park J, Woo J, Jin Y, Ryu S. Origin of the stereospecificity in binding hydroxamates of alpha- and beta-phenylalanine methylamide to thermolysin revealed by the X-ray crystallographic study. Bioorg Med Chem. 2003;11:2421-6 pubmed
    ..methylamide (1) and N-formyl-N-hydroxy-beta-phenylalanine methylamide (2) were evaluated as inhibitors for thermolysin (TLN) to find that while the D-form is more potent than its enantiomer in the case of the hydroxamate of alpha-..
  42. Carr S, Hauschka P, Biemann K. Gas chromatographic mass spectrometric sequence determination of osteocalcin, a gamma-carboxyglutamic acid-containing protein from chicken bone. J Biol Chem. 1981;256:9944-50 pubmed
    ..The structure is highly homologous to the sequence of the corresponding protein isolated from bovine bone, and the relative sequence locations of the Gla residues and the disulfide bridge are conserved. ..
  43. Breidenbach M, Brunger A. 2.3 A crystal structure of tetanus neurotoxin light chain. Biochemistry. 2005;44:7450-7 pubmed
    ..overall structure of TeNT-LC is similar to the other known CNT light chain structures, including a conserved thermolysin-like core inserted between structurally distinct amino- and carboxy-terminal regions...
  44. Sun Q, Yang Q, Gong S, Fu Q, Xiao Q. Synthesis and enzymatic evaluation of phosphoramidon and its ? anomer: Anomerization of ?-l-rhamnose triacetate upon phosphitylation. Bioorg Med Chem. 2013;21:6778-87 pubmed publisher
    ..The determined Ki and Kd values establish that phosphoramidon prepared by this method possesses excellent biological activity, and indicate that the contacts of rhamnose moiety with the enzyme have limited contribution to the binding. ..
  45. Youn K, Park C. Investigating the effect of temperature on transient partial unfolding by proteolysis. Protein Pept Lett. 2009;16:1093-7 pubmed
    ..E. coli ribonuclease H (RNase H) has been shown to be cleaved by thermolysin at the amide bond between Thr92 and Ala93 through partial unfolding...
  46. Rajan S, Yang X, Shuvalova L, Collart F, Anderson W. YfiT from Bacillus subtilis is a probable metal-dependent hydrolase with an unusual four-helix bundle topology. Biochemistry. 2004;43:15472-9 pubmed
    ..of the metal and the residues that constitute the putative active site in YfiT are similar to those of metalloproteases such as thermolysin. Our structural analyses suggest that YfiT might function as a metal-dependent hydrolase.
  47. Xiang H, Xiang G, Lu Z, Guo L, Eckstein H. Total enzymatic synthesis of cholecystokinin CCK-5. Amino Acids. 2004;27:101-5 pubmed
    ..Immobilized enzymes were used for the formation of all peptide bonds except thermolysin. Beginning the synthesis with phenylacetyl (PhAc) glycine carboxamidomethyl ester (OCam) and H-Trp-OMe by using ..
  48. DeCaluwé G, Degrip W. Point mutations in bovine opsin can be classified in four groups with respect to their effect on the biosynthetic pathway of opsin. Biochem J. 1996;320 ( Pt 3):807-15 pubmed
    ..Taking any functional consequences of these mutations into consideration, it seems that point mutations can have mosaic effects and therefore should be examined at several levels (folding, targeting, functional parameters). ..
  49. Ploug M, Kjalke M, Rønne E, Weidle U, Høyer Hansen G, Danø K. Localization of the disulfide bonds in the NH2-terminal domain of the cellular receptor for human urokinase-type plasminogen activator. A domain structure belonging to a novel superfamily of glycolipid-anchored membrane proteins. J Biol Chem. 1993;268:17539-46 pubmed
    ..amino acid composition, and sequence analyses of peptides generated by trypsin, endoproteinase Asp-N, and thermolysin. The following disulfide bond structure was determined: Cys3-Cys24, Cys6-Cys12, Cys17-Cys45, and Cys71-Cys76...
  50. Bowler M, Guijarro M, Petitdemange S, Baker I, Svensson O, Burghammer M, et al. Diffraction cartography: applying microbeams to macromolecular crystallography sample evaluation and data collection. Acta Crystallogr D Biol Crystallogr. 2010;66:855-64 pubmed publisher
    ..The implementation of these techniques in the context of planned upgrades to the ESRF's structural biology beamlines is also presented. ..
  51. Van Quynh A, Willson S, Bryant R. Protein reorientation and bound water molecules measured by 1H magnetic spin-lattice relaxation. Biophys J. 2003;84:558-63 pubmed
  52. Lung S, Chuong S. A transit peptide-like sorting signal at the C terminus directs the Bienertia sinuspersici preprotein receptor Toc159 to the chloroplast outer membrane. Plant Cell. 2012;24:1560-78 pubmed publisher
  53. Sjøli S, Nuti E, Camodeca C, Bilto I, Rossello A, Winberg J, et al. Synthesis, experimental evaluation and molecular modelling of hydroxamate derivatives as zinc metalloproteinase inhibitors. Eur J Med Chem. 2016;108:141-153 pubmed publisher
    ..synthesised a series of hydroxamate derivatives and validated the compounds as inhibitors of the M4 enzymes thermolysin and pseudolysin, and the endogenous metalloproteinases ADAM-17, MMP-2 and MMP-9 using experimental binding ..